Term
-__ is the process in which the DNA within a cell makes an exact copy of itself.
-- __: cell cycle arrested
-- __: cellular contents, excluding the chromosomes are duplicated
-- __: Each of the 46 chromosomes is duplicated by the cell
-- __: the cell double checks the duplicated chromosomes for error, making any needed repairs |
|
Definition
- dna replication
-- G0
-- G1
-- S
-- G2 |
|
|
Term
|
Definition
|
|
Term
- dna replication in ecoli requires more than __
- five of these are __, which catalyze the synthesis of new dna
- __ and __ play key roles in dna replication |
|
Definition
- 20 proteins
- dna polymerases
- poly 3; poly 1 |
|
|
Term
- polymerase I: __ removal and dna __; __ and __ exonuclease
- polymerase II: function is __, __ exonuclease, attachment of __ to __
- polymerase III: fuction is __, __ exonuclease
- polymerase IV: function is __; attachement of __ to __
- polymerase V: function is __, attachment of __ to __, __, and __ |
|
Definition
-primer; repair; 5>3; 3>5
- repair; 3>5; bulky hydrocarbons; bases
- replication; 3>5
- repair; bulky hydrocarbons; bases
- repair; bulky hydro carbons; bases; sites missing bases; covalently joined bases |
|
|
Term
| polymerases __, __, and __ can replicate through regions of damaged DNA. they are called __ polymerases or error-prone polymerases |
|
Definition
|
|
Term
| __ catalyzes __ bond formation |
|
Definition
| dna polymerase; phosphodiester |
|
|
Term
important characteristics of dna synthesis
1. __ and __ are required
-- these include __, __, __, __
2. a __ is used to direct dna synthesis
-- __ is a __ enzyme
3. a __ from which the new strand grows must be present |
|
Definition
1. deoxynucleoside triphosphates; mg+2
-- dATP; dGTP; dCTP; dTTP
2. template strand
-- dna polymerase; template directed
3. primer |
|
|
Term
chain elongation is a __ reaction
- base pair between incoming __ and a residue of the __
- terminal __ attacks __ of incoming nucleotide to form a new __ linkage
- nucleotides added one at a time to the __ end
- chain grown is always __ |
|
Definition
nucleotidyl group transfer
- deoxynucleotide 5' triphosphate; parental strand
- 3' OH; alpha phosphorous; phosphodiester
- 3'
- 5>3 |
|
|
Term
important characteristics of dna synthesis
4. many __ have nuclease activity that allows for the removal of mismatched bases
- __ activity contributes to the remarkably high fidelity of dna replication, less than 10^-8 errors per base pair
- most uncorrected replication errors are subsequently corrected by the __ |
|
Definition
4. dna polymerases
- 3>5 exonuclease
- dna repair enzymes |
|
|
Term
| proofreading by the __ activity of __ during dna replication |
|
Definition
| 3>5 exonuclease; dna polymerases |
|
|
Term
dna polymerase structure
- the structures of dna polymerases are __
- the region containing the active site approximates the shape of a __
- the __ and __ domains wrap around the dna positioning it in the active site located in the __ domain |
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Definition
- similar
- right hand
- thumb; fingers; palm |
|
|
Term
-Specificity of Replication is Dictated by the __ of __
-The specificity of replication is determined not only by the correct __ between the incoming dNTP and the DNA template, but also by the overall __ of the incoming base.
- an analog of adenosine that cannot form h bonds still directs incorporation of __ |
|
Definition
- complementarity; bases
- hydrogen bonding; shape
- t |
|
|
Term
__ Contributes to Fidelity
-Binding of the incoming __ induces a structural change in which the __ domain closes over the nucleotide, forming a pocket into which only the correct nucleotide fits. |
|
Definition
induced fit
- dNTP; finger |
|
|
Term
-__, powered by __, separate the strands of the double helix to make the DNA available for __.
-The __, which consists of a ring-like structure composed of __ subunits, acts as a wedge to pry the helix apart. |
|
Definition
-helicases; atp hydrolysis; dna polymerase
- helicase; six |
|
|
Term
| Defects in the helicase can result in __, a pathological condition characterized by premature aging. |
|
Definition
|
|
Term
__ Prepare the Double Helix for Unwinding
-When DNA is unwound for replication, -__ occurs because of overwinding in nearby regions of the helix.
-__ induce or eliminate supercoils.
-__ relax supercoils while __, such as DNA gyrase in E. coli, introduce supercoiling. |
|
Definition
topoisomerases
- supercoiling
- topoisomerases
- type I topoisomerases; type 2 topoisomerases |
|
|
Term
__ is a Therapeutic Target
-__ and __ are inhibitors of DNA gyrase that are used to treat bacterial infections.
-__ prevents anthrax poisoning |
|
Definition
bacterial topoisomerase
- nalidixic acid; ciprofloxacin
- ciprofloxacin (cipro) |
|
|
Term
| dna replication is highly __ and very __ |
|
Definition
|
|
Term
e coli dna replication begin at a unique site
-Replication begins in E. coli at a unique 245 base pair site called the __.
-__ protein binds at distinct sites in the locus and then recruits __, a helicase that uses ATP hydrolysis to unwind the locus. |
|
Definition
|
|
Term
e coli dna replication begin at a unique site
-__ bind to the __-rich sites in the locus to prevent reannealing of the helix.
-The resulting complex of DNA and proteins is called the __ |
|
Definition
- single stranded binding protein (ssb); AT
- prepriming complex |
|
|
Term
- dna polymerase cannot initiate new __
--unable to covalently link 2 individual __ together
-- able to covalently link a __ to a __ |
|
Definition
- strands
-- nucleotides
-- nucleotide; primer |
|
|
Term
__ Synthesized by __ Enables DNA Synthesis to Begin
-A special type of __ called __ synthesizes short RNA (≈10 nucleotides) complementary to a DNA strand that then serves as a __ for DNA synthesis. |
|
Definition
Rna primer; primase
- rna polymerase; primase; primer |
|
|
Term
-Primase is part of a large complex called the __
-__, employing a third enzymatic activity, a __ activity, subsequently removes the primers and replaces the RNA with __. |
|
Definition
- primasome
- dna polymerase I; 5>3 exonuclease; dna |
|
|
Term
One Strand of DNA is Made __ while the Other Strand is Made in __
-The site of replication is called the __.
-The fork moves in one direction, so both strands are copied __.
-However, all DNA polymerases synthesize DNA only in the __ direction. |
|
Definition
continuously; fragments
- replication fork
- simultaneously
- 5>3 |
|
|
Term
-At the replication fork, the __ strand is synthesized continuously in the __ direction.
-The __ strand is synthesized discontinuously as small pieces called __ in the __ direction. |
|
Definition
- leading; 5>3
- lagging; okazaki fragments; 5>3 |
|
|
Term
__ begins each okazaki fragment
-__ is a complex containing __ enzyme which synthesizes short pieces of RNA at the replication fork (complementary to the lagging-strand template)
-__ uses the __ to start the lagging-strand DNA synthesis |
|
Definition
rna primer
- primosome; primase
- dna polymerase; rna primer |
|
|
Term
- The holoenzyme responsible for the majority of DNA synthesis in E. coli is __, a multi-protein complex.
-__ is highly processive, once it begins catalysis it rarely releases the __. |
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Definition
- dna polymerase III
- dna polymerase III; dna substrate |
|
|
Term
-The source of the processivity is the __, a dimer that encircles the helix.
-The sliding clamp is loading onto the DNA by a protein called the __, which uses the energy of __ to open and close the clamp |
|
Definition
- sliding clamp (Beta 2 subunit)
- clamp loader; atp hydrolysis |
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Term
-The __ strand is looped so that it passes through the polymerase active site in the __ direction, allowing synthesis to occur in the __ direction
-__ binds the single stranded DNA - replication does not result in large patches of open __. |
|
Definition
- lagging;3>5; 5>3
- ssb; ssDNA |
|
|
Term
as replication proceeds:
-__ unwinds DNA template
-replication continues on __ strand
-__ makes a new primer on the __ strand
-__ strand core __ finishes an __ |
|
Definition
- helicase
- leading
- primase; lagging
- lagging; polymerase; okasaki fragment |
|
|
Term
-Lagging strand polymerase encounters preceding Okasaki fragment, releases the lagging strand. Nick between Okasaki fragments will be sealed by __ and __
-Polymerase core on __ strand never releases - keeps replication processive |
|
Definition
- polymerase I; dna ligase
- leading |
|
|
Term
| Lagging strand polymerase then binds newly made __ - starts next Okasaki fragment |
|
Definition
|
|
Term
Okazaki Fragments Are Joined by Action of __ and __
-Okazaki fragments are joined to produce a continuous strand of DNA in 3 steps:
(1) Removal of the __ by The __ activity of __
(2) Synthesis of replacement DNA, __: polymerase activity of pol I synthesizes DNA in place of RNA
(3) Sealing of adjacent DNA fragments by __ |
|
Definition
dna poly I; dna ligase
- rna primer; 5>3 exonuclease; dna pol 1
- nick translation
- dna ligase |
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Term
Problem: after Okazaki fragment synthesis is complete, have gap between end of Okasaki fragment and RNA primer
-__ extends the Okasaki fragment while its __ activity removes the __. This is called __.
-__ then dissociates, __ binds to the nick.
-__ then catalyzes the formation of a __ linkage between 3’-hydroxyl and 5’-phosphate of adjacent Okazaki fragments - enzyme then dissociates from DNA |
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Definition
- dna pol I; 5>3 exonuclease; rna primer; nick translation
- dna pol I; dna ligase
- dna ligase; phosphodiester |
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|
Term
-__ chromosomes are large linear, double-stranded DNA molecules
-Rate of __ slower than in E. coli but multiple __ enables rapid replication of genome |
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Definition
- eukaryotic
- replication; origins |
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|
Term
-Multiple __ of replication are required with each origin of replication representing a replication unit or replicon. __ have ≈ 30,000 replicons.
-Proteins called __ allow only one replication per replicon per round of DNA synthesis.
-__, which displays primase activity, initiates DNA replication in eukaryotes, generating a DNA molecule ≈ 20 dNTP in length.
-__, a more processive enzyme, extends the chains. The switch from polymerase __ to __ is called polymerase switching. |
|
Definition
- origins; humans
- licensing factors
- dna polymerase alpha
- dna polymerase delta; alpha; delta |
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|
Term
| __: unique structures at the ends of linear chromosomes |
|
Definition
|
|
Term
__, the free end of linear dna, have two problems
1. they are susceptible to damage by __
2. due to the nature of dna synthesis, one strand will __ upon each round of dna synthesis |
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Definition
telomeres
1. nucleases
2. shorten |
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Term
__: protecting the end from degradation
- the end of the __ strand is rich in __, and contains hundreds of tandem repeats
- the stretch of __ rich single strand dna can form a loop structure to protect the end of a chromosome |
|
Definition
telomeres
- leading; guanine
- guanine |
|
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Term
__ replicates telomeres
-__ contains an __ that it uses to extend the leading strand.
-This prevents the lagging strand from getting progressively shorter
-In rapidly dividing cells, including __, telomeres must be maintained by the telomerase to prevent shortening of the __, which would lead to cell death.
-High __ activity is a characteristic of cancer cells |
|
Definition
telomerase
- telomerase; rna template
-
- cancer cells; lagging
- telomerase |
|
|
Term
__: an enzyme that can use RNA as a template to synthesize DNA
-present in retroviruses, such as __
-also called __
-high error rate because it has no __ capabilities
-target of __ drugs |
|
Definition
reverse transcriptase
- hiv
- rna directed dna polymerase
- proof-reading
- antiviral |
|
|
Term
-DNA is the only cellular macromolecule that can be __ - __ can have a very high cost
-DNA damage includes: __ modifications, __ deltions or insertions, __ of dna strands, breakage of __
-DNA damage can result from __ (chemicals, radiation) and from __ made during normal cellular processes |
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Definition
- repaired; mutations
- base; nucleotide; cross linking; phosphodiester bonds
- environmental agents; errors |
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|
Term
-The simplest source of DNA damage is the incorporation of an incorrect base during __ that escapes proofreading by the __.
-Other errors include insertions, deletions or breaks in one or both strands, which may halt DNA synthesis altogether.
-Special DNA polymerases, called __, can replicate across the damage and generate a rough draft of the damaged sequence. These are not as precise as replicative polymerases |
|
Definition
- replication; dna polymerases
-
- translesion polymerases |
|
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Term
-Some sequences, such as __ regions, are inherently prone to errors
-Some genetic diseases such as Huntington’s Disease are related to expansion of these regions. __ is repeated up to 31 times in unaffected people, much more in those with the disease
-Formation of alternative structures during replication leads to the expansion |
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Definition
- triplet repeat
- huntington's disease; CAG (encoding glutamine) |
|
|
Term
- bases can be damaged by __, __, and __
-Chemicals that alter specific bases after replication is complete are called __.
-Hydroxyl radicals oxidize __ to __, which base pairs with __ instead of __ during the next round of replication. |
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Definition
-oxidizing agents; alkylating agents; light
-mutagens
- guanine; 8-oxoguanine; adenine; cytosine |
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|
Term
| -Deamination can be mutagenic. __ can be deaminated, forming __, which pairs with __ instead of __ |
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Definition
| - adenine; hypoxanthine; cytosine; thymidine |
|
|
Term
-Bases can react with hydrocarbons in an __ reaction.
-__ is converted into a highly reactive epoxide that reacts with __ forming a compound that, during replication, converts a __ base pair into a __ base pair.
-Chemicals in cigarette smoke and automobile exhaust can also be converted into reactive __
-Ultraviolet radiation covalently links adjacent __ thereby blocking replication. |
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Definition
- alkylation reaction
- aflatoxin; guanine; g-c; a-t
- epoxides
- pyrimidines |
|
|
Term
DNA repair systems follow the same mechanistic outline:
1.__ the inappropriate base(s).
2.__ the inappropriate base(s).
3.Fill in the resulting gap with DNA __ and __.
-The first DNA repair mechanism occurs when __ proofreads the newly synthesized DNA and correct mismatches. |
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Definition
1. recognize
2. remove
3. dna poly I; dna ligase
- dna polymerase |
|
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Term
-Some damage can be corrected with direct repair
-T-T dimers can be directly repaired by many organisms (but not __ or other placental mammals)
-Repair of thymine dimers by __ - __ |
|
Definition
-
- humans
- dna photolyase; photoreactivation |
|
|
Term
-__ correct errors not corrected by proofreading that cannot be directly repaired
-In E. coli, need one protein to recognize the error (__), one to recruit an exonuclease to cleave the DNA (__), and the exonuclease (__).
-Parent strand may be “marked” by __ of adenines so that repair machinery can recognize the correct base |
|
Definition
- mismatch repair systems
- mutS; MutL; MutH
- methylation |
|
|
Term
-__ uses several enzymes to correct modified bases.
-__ such as AlkA remove the base. The site of the missing base is called an __.
-The DNA backbone at the AP site is cleaved by __ and a __ excises the deoxyribose phosphate.
-The gap is repaired by __ and __ |
|
Definition
- base-excision repair
- glycosylases; ap site
- ap endonuclease; phosphodiesterase
- dna poly I; dna ligase |
|
|
Term
-Hydrolytic deamination of __ to __ is one of most common types of DNA damage
-Uracil in place of cytosine causes incorporation of an incorrect base during replication
-Corrected by __
-__ hydrolyze base-sugar N-glycosidic bonds
-__ bases are then removed and replaced |
|
Definition
- cytosine; uracil
-
- base excision repair
- dna glycosylases
- deaminated |
|
|
Term
-Use of __ in DNA preserves the integrity of the genetic information.
-If uracil naturally occurred as a base in DNA, __ would lead an A-U pair replacing a G-C pair after the next round of replication.
-Use of __ instead of __ allows the detection of deamination of __.
-If uracil is detected in DNA, it is removed by __ and the resulting AP is repaired with the insertion of __. |
|
Definition
- thymidine
- cytosine deamination
- thymidine; uracil; cytosine
- uracil dna glycosylase; cytosine |
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|
Term
-If base-excision fails to recognize the damaged base, the mutation may be corrected by the __ system.
-An enzyme complex recognizes the distortion of the DNA caused by offending base. The __ cleaves the DNA at two sites, several nucleotides on each side of the distortion.
-__ and __ close the resulting gap |
|
Definition
- nucleotide-excision repair
- UvrABC excinuclease
- dna poly I; dna ligase |
|
|
Term
what happens when excision repair does not function?
-__– genetic disorder in DNA nucleotide excision repair genes – a number of different proteins involved
-Patients have extreme sensitivity to the sun, much greater risk of skin cancer, and about 20% have neurological disorders |
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Definition
| - xeroderma pigmentosum (xp) |
|
|
Term
-__ are caused by mutations in genes that control growth.
-__ act a tumor suppressors.
-If both copies of a DNA repair enzyme are mutated, __ is more likely to develop
-Because tumors lack DNA repair systems, damaging the DNA with chemicals such as __ and __ is a strategy to prevent cancer growth. |
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Definition
-cancers
- dna repair enzymes
- cancer
- cyclophosphamide; cisplatin |
|
|
Term
-The __ test - simple, sensitive means of determining if a chemical is mutagenic.
-Example: bacteria that require histidine because of a mutation in its biosynthetic pathway are treated with the chemicals. Then grow the bacteria without the required amino acid. Bacterial growth (appearance of colonies) suggest that the chemical caused a mutation which restored histidine synthesis |
|
Definition
|
|
Term
-__ - exchange or transfer of pieces of DNA from one chromosome to another or within a chromosome
-__ - occurs between pieces of DNA that have closely related sequences (e.g. exchange between paired chromosomes during meiosis). Genetic recombination is a type of homologous recombination.
-__ occurs between unrelated sequences (e.g. Transposons )
-__ - recombination at a specific location |
|
Definition
-recombination
- homologous recombination
- nonhomologous recombination
- site specific recombination |
|
|
Term
why is recombination needed?
-__ gives new genetic variation
- __ creates different combinations of mutations within a genome - parents chromosomes are not the same as progeny chromosomes
of DNA |
|
Definition
- mutation
- recombination |
|
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Term
-__ probably evolved to help DNA repair (confers a selective advantage)
-Severe __ in DNA can be repaired by strand exchange from intact daughter chromosome to defective daughter
-Very important process - more than 100 genes from E. coli are involved in recombination and repair |
|
Definition
-recombination enzymes
- lesions |
|
|
Term
Recombination is a complicated process – many proteins!
1.Double strand break is recognized,the __ ends are digested, single-strand regions are bound by __.
2.__ occurs - single strand from damaged DNA replaces a strand in undamaged DNA. This three-stranded structure is called a __.
3.__ occurs, using the undamaged DNA as a template.
4.A second strand invasion occurs to complete the repair, forming a structure called a __.
5.Cleavage and ligation yield two intact __ |
|
Definition
1. 5'; rad51
2. strand invasion; displacement loop
3. dna synthesis
4. holliday junction
5. helices |
|
|
Term
what happens when ds break repair doesn't function?
-__ and __ both required for normal recombinational repair of ds breaks in DNA
-form complex with __ (eukaryotic equivalent of RecA)
-approx. 12,000 cases of breast cancer a year in north America are related to mutations in these two proteins |
|
Definition
|
|
Term
-BRC repeats of __ interact directly with __
-__ may serve to deliver __ to the site of DNA damage
-Conserved __ appears to bind ssDNA and dsDNA |
|
Definition
-brca2; rad51
- brca2; rad51
- c-terminus |
|
|
Term
| The synthesis of RNA from a DNA template is called transcription, a process catalyzed by __ |
|
Definition
|
|
Term
RNA polymerase has the following requirements:
1.A __. The sequence of the newly synthesized RNA is complementary to the DNA template. The DNA strand that has the same sequence as the RNA product (with T instead of U) is called the __.
Activated precursors in the form of the
2.four __
3.Divalent metal ions, usually __ or __ |
|
Definition
1. template; coding strand
2. ribonucleoside triphosphates (atp, gtp, utp, ctp)
3. Mg+2; Mn+2 |
|
|
Term
| RNA polymerase initiates and elongates the RNA product, with the chain growing in the __ direction. The 3’OH of the growing chain attacks the inner most __ of the incoming ribonucleoside triphosphate. |
|
Definition
| -5>3; phosphoryl (alpha) group |
|
|
Term
A __ is a transcriptional unit. Types of RNA made are:
(1) __
--Carries amino acids to translation machinery
--Very stable molecules
(2) __
--Makes up much of the ribosome
--Very stable, majority of cellular RNA
(3) __
--Encodes message from DNA to ribosomes
--Rapidly degraded by __
--Small fraction of steady-state RNA, but use a large percentage of synthetic capacity of cell
(4) __
-- participate in metabolic events; many have catalytic activity |
|
Definition
gene
1. tRNA
2. rRNA
3. mRNA
--
-- nucleases
4. small RNA |
|
|
Term
E. Coli RNA Polymerase Is an Oligomeric Protein
-Core enzyme: __ participates in many of the transcription reactions
-The holoenzyme, consisting of __ subunits, initiates RNA synthesis.
-The core enzyme, composed of ___ subunits, elongates the RNA product.
-__: role in initiation of transcription |
|
Definition
- a2bbw
-α2ββ’ωσ
-α2ββ’ω
- sigma subunit |
|
|
Term
-The three stages of RNA synthesis are __, __, and __.
-How does the RNA polymerase know where to start? Specific __ sequences called __ direct __ to the proper initiation site.
-In E. coli, two DNA sequences that act as a promoter for many genes are the __ and the __. |
|
Definition
- initiation; elongation; termination
- dna; promoters; rna polymerase
- -10 sequence (pribnow box); -35 sequence |
|
|
Term
-__ will closely match the consensus sequence
-__ tend to have multiple substitutions at these sites
-Other sequences __ of the promoter and the action of transcription factors can enhance promoter effectiveness. |
|
Definition
- strong promoters
- weak promoters
- upstream |
|
|
Term
-The __ of prokaryotic RNA polymerases is required for promoter recognition and formation of the complex
-s Subunit increases the affinity of the __ for specific __ sequences
-s subunit also decreases the affinity of the __ for __ regions
-Core polymerase (no s subunit) binds __ nonspecifically |
|
Definition
- sigma subunit
- core polymerase; promoter
- core polymerase; nonpromoter
- dna |
|
|
Term
-The s factor acts __ - one s can aid many __ in finding promoters
-E. Coli has many different s factors that recognize different consensus sequences at the promoter |
|
Definition
| - catalytically; polymerases |
|
|
Term
-__ rapidly searches for and binds promoter
- Unwinding of DNA at the initiation site requires a __ – often the rate limiting step
-RNA pol (R) and promoter (P) shift from: __ to __ |
|
Definition
- rna polymerase
- conformational change
- (RPc) closed complex (dsDNA); (RPo) open complex (17 bp unwound) |
|
|
Term
-__ begins transcription
-Once the DNA is unwound, __ can take place.
-__ can start RNA synthesis de novo – no primer needed! |
|
Definition
- rna polymerase
- elongation
- rna polymerase |
|
|
Term
- elongation takes place at __
- The region containing the RNA polymerase, DNA, and the RNA product is called the __.
-The transcription bubble moves along the DNA as DNA is unwound and then rewound, while the RNA product is extruded from the complex. A __ helix of approximately __ is an intermediate in RNA synthesis |
|
Definition
- transcription bubbles
- transcription bubble
- dna-rna hybrid helix; 8 nucleotides |
|
|
Term
-Transcription continues until a __ is reached
-Transcription complexes disassemble at the __ end of genes at specific __
-Two types of termination sequences
1) __ - generally associated with a region of the RNA transcript that forms a hairpin structure, with a series of __ |
|
Definition
- termination signal
- 3'; termination sequences
-
1) unstable elongation complex; u residues |
|
|
Term
-__ pauses after it has synthesized RNA that folds into a __
-__ with __ is unstable – __ is the weakest of the Watson-Crick base pairs
-Pause caused by __ permits weak RNA-DNA hybrid to __ |
|
Definition
- rna polymerase; hairpin
- RNA-DNA hybrid; oligo(u); rU-dA
- hairpin; dissociate |
|
|
Term
- __ dependent termination sites
-__ triggers disassembly of the transcription complex at some pause sites
-__ binds to __ chain, destabilizing the RNA-DNA hybrid and terminating transcription
-A hexamer of rho binds a __ stretch on RNA
-Note that both types of termination involve __ on the newly synthesized RNA |
|
Definition
- rho
- rho
- rho; ssRNA
- 72 nt
- signals |
|
|
Term
-Precursors of __ and __ are cleaved and chemically modified after transcription
-Although __ undergoes little or no modification after synthesis in bacteria, __ or __ are modified:
1. The final mature __ is cleaved from a larger precursor molecule.
2.Many __ transcripts lack __ sequence at the __ end of the strand. These nucleotides are added post-transcriptionally.
3.The bases and riboses of tRNA and rRNA are modified, for instance, by the attachment of __. |
|
Definition
- tRNA; rRNA
- mRNA; rRNA; tRNA
1. RNA
2. tRNA; CCA; 3'
3. methyl groups |
|
|
Term
-The antibiotics __ and __ inhibit transcription, but by different mechanisms.
-__ inhibits initiation by binding to the polymerase and blocking the exit of the nascent RNA.
-__ intercalates between the bases of the DNA double helix, preventing the DNA from being used as a template |
|
Definition
- rifampicin; actinomycin
- rifampicin
- actinomycin |
|
|
Term
-The __, an Example of Negative and Positive Regulation
-E. coli can use __ and other __ as a carbon source when __
-Uptake and catabolism of __ requires three proteins:
1.
2.
3. |
|
Definition
- lac operon
- lactose; beta galactosides; glucose is absent
- beta-galactosides
1. lactose permease (lac Y)
2. beta galactosidase (lac Z)
3. thiogalactoside transacetylase (lac A) |
|
|
Term
(1) Lactose permease (lacY) - __ for uptake of __
(2) b-Galactosidase (lacZ) - hydrolyzes __ to __
(3) Thiogalactoside transacetylase (lacA): __ nonmetabolizable __ |
|
Definition
1. transporter; beta galactosides
2. beta galactosides; hexoses
3. acetylates; beta galactosides |
|
|
Term
-When glucose is __ these proteins are synthesized in limited amounts
-When glucose levels are __ these proteins are synthesized in larger amounts if __ are present |
|
Definition
- present
- low; beta galactosides |
|
|
Term
-Three genes - lacZ, lacY and lacA form an __ that is transcribed from a single __
-Resulting large __ molecule contains three separate protein-coding regions (3 genes)
-operons with multiple protein-coding regions are common in __ but not __, though they are used in C. elegans |
|
Definition
- operon; promoter (Plac promoter)
- mRNA
- prokaryotes; eukaryotes |
|
|
Term
- __ Blocks Transcription
-lac Operon expression is controlled by the __ - __ is encoded by the __ (under control of __), located upstream from the lac operon
-lac Repressor binds simultaneously to __
-__ is adjacent to the promoter, __ is within the coding region of lacZ |
|
Definition
- lac repressor
- lac repressor protein; lac repressor; lacl gene; (p promoter)
- 2 repressor binding sites (operators)
- operator (O1); (O2) |
|
|
Term
-The __ binds simultaneously to both operators O1 and O2
-This causes the DNA to form a __ which blocks initiation of transcription
-__ act as inducers to cause the repressor to dissociate from the operon allowing __ to continue |
|
Definition
- lac repressor
- stable loop
- beta galactosides; transcription |
|
|
Term
- __: inducer for lac operon
-Several __ can act as inducers to start gene transcription
-In the presence of lactose, __ is the inducer, binds tightly to __ and causes a conformational change that decreases affinity of lac repressor for __
-b-Galactosidase converts __ to __ |
|
Definition
- allolactose
- beta-galactosides
- allolactose; lac repressor; operator
- lactose; allolactose |
|
|
Term
-__ Regulatory Protein Activates Transcription
-The lac operon is maximally transcribed when __ are the only carbon source
-When __ is present transcription is reduced 50-fold, (catabolite repression)
-A weak __ in these operons is promoted to a stronger one by an activator (in the absence of glucose)
-cAMP regulatory (or receptor) protein (__) is the activator - __ is also known as __
-cAMP levels increase when __ |
|
Definition
- cAMP
- beta galactosides
- glucose
- promoter
- (crp); CRP; catabolite activator protein (CAP)
- glucose decreases |
|
|
Term
-Some __ can sense environmental signals as well as encode proteins.
-These mRNA have special structures, called __, that bind small molecules that cause a structural change in the __ that terminates the synthesis of the mRNA. |
|
Definition
- bacterial mRNA
- riboswitches; riboswitch |
|
|
Term
-The basics of RNA synthesis are the same for __ and __
-__ organisms use differential gene regulation to generate different cell types. |
|
Definition
- eukaryotes; prokaryotes
- multicellular |
|
|
Term
Gene expression in eukaryotes is influenced by three important characteristics:
1. __ regulation
2.__ processing, including extensive processing of __ precursors
3.The __, which separates the site of __ synthesis from that of __ synthesis |
|
Definition
1. complex transcription
2. RNA; mRNA
3. nuclear membrane; RNA; protein |
|
|
Term
- eukaryotic cells have three __
-RNA synthesis is catalyzed by three __ that differ in DNA substrate specificity, location, and sensitivity to the toxin α-amanitin.
-All the polymerases are similar in structure, but RNA polymerase II has a unique domain, called the __, that plays an important regulatory role |
|
Definition
- rna polymerases
- rna polymerases
- carboxyl terminal domain |
|
|
Term
-RNA poly I: location is __; transcripts include __, __, __; effect on alpha amanitin: __
-RNA poly II: location is __; transcripts include __ and __; effect on alpha amanitin: __
-RNA poly III: location is __; transcripts include __ and __; effect on alpha amanitin: __ |
|
Definition
- nucleolus; 18s, 5.8s, 28sRNA; insensitive
- nucleoplasm; mRNA and snRNA; strongly inhibited
- nucleoplasm; tRNA and 5sRNA; inhibited by high [] |
|
|
Term
-Eukaryotic promoters, also called __, are more complicated than bacterial promoters. Each type of polymerase has distinct promoters.
-The promoters bind to proteins, called __ or __, that regulate polymerase activity. |
|
Definition
- cis-acting elements
- trans-acting elements; transcription factors |
|
|
Term
RNA polymerase II
-Transcribes all __ genes and some small __ encoding genes
-Protein-encoding RNA synthesized by RNA pol II is called __
-20%-40% of all cellular __
-General ___ interact directly with RNA pol II and control initiation at class II genes
-Elaborate regulation of RNA pol II accounts for cell differentiation and development. |
|
Definition
- protein-encoding; rna
- mRNA precursor (or hnRNA)
- rna synthesis
- transcription factors |
|
|
Term
-Most genes transcribed by RNA pol II have an A/T rich sequence called __ located -24 to -32
-Other important sequences include the __ at the transcriptional start site and a __ at +28 to +32
-Many promoters also have __ or __ boxes upstream |
|
Definition
- tata box
- initiator element; downstream core promoter element
- caat; gc |
|
|
Term
-The __ complex initiates assembly of the active transcription complex
- Transcription factors of RNA poly II= __
-In genes with a TATA box, the __, a component of TFIID, binds to the TATA box. Other transcription factors bind to generate the basal transcription complex.
-__ binds to the complex and phosphorylates the __ of the polymerase, facilitating the transition from initiation phase to the elongation phase. |
|
Definition
- TFIID
- TFII
- tata binding protein (TBP)
- TFIIH; carboxyl terminal domain |
|
|
Term
-Enhancer sequences are __ that have no promoter activity but can stimulate the effectiveness of promoters even when located thousands of nucleotides from the start site.
-Inappropriate enhancer use may cause __
-Chromosomal translocations can place a gene under the control of a powerful enhancer. For instance dysregulation of the gene myc, a transcription factor, resulting from the translocation of an enhancer to a region near the myc gene plays a role in the development of __ and __ |
|
Definition
- cis-acting elements
- cancer
- burkitts lymphoma; b-cell leukemia |
|
|
Term
-Many other proteins play a role in regulating the efficiency and specificity of gene transcription
-A large complex called __ acts as a bridge between enhancer-bound activators and proteins, including the polymerase, at the promoter.
-Effects on transcription can
depend on the other components
of the regulatory complex
(combinatorial control). |
|
Definition
|
|
Term
- gene expression is regulated by __
-Gene expression in eukaryotes must respond both to conditions within the __ and to __
-__ hormones are one class of regulatory molecules that control gene expression.
-__ controls the genes in the development of female secondary sex characteristics. It exerts its effects by forming a complex with a specific receptor protein called the estrogen receptor. |
|
Definition
- hormones
- cell; external stimuli
- steroid
- estradiol |
|
|
Term
-__ have similar domain structures
-The estrogen receptor is part of a larger class of regulatory proteins called __, all of which are activated by binding of small molecules or ligands.
-__ (previous) bind to specific regions of the DNA called __. Thus, the estrogen receptor binds to the estradiol response element (ERE). |
|
Definition
- nuclear hormone receptors
- nuclear hormone receptors
- nuclear hormone receptors; response elements |
|
|
Term
Nuclear hormone receptors have two conserved domains.
1.The __ has zinc-finger domains that confer specific DNA binding.
2.Ligand binding to the
__
causes a structural change
that enables the receptor
to recruit other proteins to
regulate transcription. |
|
Definition
1. dna binding domain
2. ligand binding domain |
|
|
Term
-__ leads to structural rearrangements in the receptor.
-These changes do NOT affect binding to the __
-Instead, the changes
promote recruitment of
additional proteins that
assist in __ |
|
Definition
- ligand binding
- response element
- transcription |
|
|
Term
- __ and __ are recruited by nuclear hormone receptors
-Proteins that bind the receptor after ligand binding are called __.
-Some nuclear hormone receptors repress transcription in the absence of ligand. __ proteins bind to them when ligand is absent to assist in repression of transcription. |
|
Definition
- coactivators; corepressors
- coactivators
- corepressor |
|
|
Term
- __ receptors are targets for drugs
-Ligands that activate a nuclear hormone receptor are called __, while ligands that inhibit the receptor are called __.
-Some __ require the estradiol-receptor complex, and growth of these cancers can be slowed by administering receptor antagonists, such as __ and __
-Such antagonists to the
estrogen receptor are
called __. |
|
Definition
- steroid hormones
- agonists; antagonists
- cancers; tamoxifen; raloxifene
- selective estrogen receptors modulators (SERMS) |
|
|
Term
-__ results in chromatin remodeling
-How do coactivators stimulate transcription? One mechanism involves loosening the interaction between histones and DNA, so the DNA is more accessible to transcription machinery.
-A common means of weakening the interaction of the histones with the DNA is by __ of the __ on specific __ residues. |
|
Definition
- histone acetylation
-
- acetylation; histones; lysine |
|
|
Term
-__ is also important for transcription!
-__ located in the nucleus generates acetyl CoA that is used by __ to modify histones.
-__ are components of
coactivators or are
recruited by coactivators.
-The __ are
extended and fit in the HAT
active site. |
|
Definition
- acetyl CoA
- atp-citrate lyase; histone acetyltransferases (HATS)
- HATs
- histone tails |
|
|
Term
1. Acetylation reduces the affinity of __ for __.
2. __ residues are interaction sites for many proteins which regulate transcription, including factors which bind the __
3. __ residues also bind and recruit chromatin remodeling proteins. |
|
Definition
1. histones; dna
2. histone acetyllysine; tata-box
3. acetyllysine |
|
|
Term
-The __ of histones is reversible, allowing genes to be expressed or repressed at the appropriate time.
-Histone __ catalyze the removal of acetyl groups, leading to __
-Histones can also be __ or __
-All covalent
modifications
of histone are
reversible |
|
Definition
- acetylation
- deacetylases; inhibition of transcription
- methylated; phosphorylated |
|
|
Term
RNA processing in eukaryotes
-Large precursor RNA (__) is processed to __, __, and __, which are components of the ribosome.
-Before cleavage, some bases and riboses are modified in a process catalyzed by RNA-protein complexes called __ |
|
Definition
- (45s); 18s; 28s; 5.8S rRNAs
- small nucleolar ribonucleoproteins (snoRNP) |
|
|
Term
Transfer RNA is extensively processed
-__ removes nucleotides from the 5’ end,while the __ adds nucleotides to the 3’ end.
-Bases and riboses are also __.
-Many eukaryotic tRNA
precursors contain an
__ that is removed by
an endonuclease, and
the resulting products
are joined by a ligase |
|
Definition
- RNase P; CCA adding enzyme
- modified
- intron |
|
|
Term
-All eukaryotic __ undergo modifications to increase their stability and make them better substrates for translation
-Ends are modified so they are no longer susceptible to __
-The __ ends are modified (capped) before the mRNA precursors are completely synthesized
-Addition of __ required for splicing and other processing events. __ is also site where ribosomes bind. |
|
Definition
- mRNA precursors
- exonuclease degradation
- 5'
- cap; cap |
|
|
Term
-Formation of a cap at the __ end of a eukaryotic mRNA precursor involves an unusual __
1. __ removes phosphate at 5' end
2. 5' end gets __ group from __, catalyzed by __, product called a cap |
|
Definition
- 5'; 5'-5' triphosphate linkage
1. phosphohydrolase
2. gmp; gtp; guanalyltransferase |
|
|
Term
- the caps can also undergo __
3. __ base is methylated
4. __ groups may also be methylated |
|
Definition
- methylation
3. n-7
4. 2'-hydroxyl |
|
|
Term
- __ at the 3' end of mRNA precursors
-Eukaryotic mRNA precursors are also modified at their 3’ ends - __ adds up to 250 adenylate residues to the 3’ end
-This poly A tail is progressively shortened by __
-The __ increases the time required for
nucleases to reach the coding region |
|
Definition
- poly a tails
- poly a polymerase
- 3' exonucleases
- poly a tail |
|
|
Term
-Most genes in eukaryotes consist of __ (coding regions) and __ (noncoding regions). Introns vary in length from 50 to 10,000 nucleotides.
-The __ are joined by splicing |
|
Definition
|
|
Term
intron-exon junctions have several common features:
1.The __ end of the junction has the sequence 5’AGGUAAGU 3’ with the first __ from the 5’ end marking the beginning of the __.
2.The 3’ end of the intron is marked by a stretch of __ followed by any base, a C, and then the intron ends with __.
3.The branch site is located 20-50 nucleotides from the __ end of the intron. |
|
Definition
1. 5'; GU; intron
2. pyrimidines (polypyrimidine tract); ag
3. 3' |
|
|
Term
-Splicing is facilitated by __ which associate with the pre-mRNA to form the splicing apparatus called the __.
-__ at the 5’ splice site, followed by __ binding at the branch site.
-Binding of __ completes splicesome formation. |
|
Definition
- snRNPs; spliceosome
- u1 snRNP; u2 snRNP
- u4-u5-u6 tri-snRNP |
|
|
Term
-The __ and the __ are the actual catalysts of splicing.
-The catalytic snRNAs facilitate __ reactions that remove the introns and join the exons. |
|
Definition
- u2 snRNA; u6 snRNA
- transesterification |
|
|
Term
-__ in either pre-mRNA or splicing factors can result in pathological conditions.
-Defects in splicing or alternative splicing may cause up to 15% of all genetic diseases.
-__, a
disease of acquired
blindness, is due to a
mutation in the
U4-U5-U6 tri-snRNP. |
|
Definition
- mutations
-
- retinitis pigmentosa |
|
|
Term
| Some __, diseases resulting from defective hemoglobin synthesis, are caused by mutations at splice sites in the pre-mRNA for the β chain of hemoglobin. |
|
Definition
|
|
Term
-Introns allow formation of alternative forms of a protein called __
-__ is a powerful mechanism for expanding protein diversity.
-In alternative splicing, a __ can be spliced in different patterns, generating proteins with different functions. |
|
Definition
- splice variants
- alternative splicing
- pre-mRNA |
|
|
Term
what controls how splicing sites are selected
-__ splicing factors bind to __ sequences in the pre-mRNA
-__ leads to different coding sequences, and proteins with different functions |
|
Definition
- trans-acting; cis acting
- alternative splicing |
|
|
Term
-Transcription and splicing are coordinated by the __ of RNA polymerase II.
Functions of the CTD include:
1.Recruiting __ to synthesize the 5’ cap.
2.Recruiting components of the __
3.Recruiting an __ that cleaves the pre-mRNA to expose the site for __. |
|
Definition
- carboxyl-terminal domain (CTD)
1. enzymes
2. splicing complex
3. endonuclease; poly a addition |
|
|
Term
-The coding information of some mRNAs can be altered by __.
-The unedited transcript forapolipoprotein B, a protein important in lipid transport, encodes a 512 kd protein,apo B-100.
-RNA editing deaminates a cytidine in the codon for glutamine forming uridine, which generates a stop codon.The edited transcript encodes for apo B-48, a 240 kd protein |
|
Definition
|
|
Term
-Some RNAs, called __, function as catalysts.
-Introns from certain organisms can self-splice. Self-splicing or __ were initially identified in rRNA from Tetrahymena.
-Group 1 introns
require __
as a cofactor. |
|
Definition
- ribozymes
- group 1 introns
- guanosine |
|
|
Term
the genetic code
-__ – four letter nucleotide “alphabet” is translated into the 20-letter amino acid “alphabet”
-__ – three nucleotides which specify an amino acid. A one letter or two letter genetic code made from four bases would not have been sufficient to encode for 20 amino acids.
-__ - each potential starting point for interpreting the 3 letter code. A nonoverlapping code is used, so the reading frame is very important
-__ - the sequence of bases is read sequentially from a fixed starting point until termination in a 5’ to 3’ direction. |
|
Definition
- translation
- codons
- reading frame
- directionality |
|
|
Term
-the genetic code is a __ code
- translation of the correct message requires selection of the correct __ |
|
Definition
-nonoverlapping
- reading frame |
|
|
Term
-almost all living organisms use the same genetic code
1.The genetic code is __. In any organism each codon corresponds to only one amino acid.
2. There are multiple codons for most amino acids (code is __), and synonymous codons specify the same __
3. The first two nucleotides of a codon are often enough to specify a given __
4. codons with similar sequences specify similar __
5. only __ of the 64 codons specify amino acids |
|
Definition
1. unambiguous
2. (degenerate); synonymous codons
3. amino acid
4. amino acids
5. 61 |
|
|
Term
- the termination codons are as follows:
- the initiation codon is __, specifies the amino acid __, and is the initiation site for protein synthesis |
|
Definition
- uaa, uga, uag
- aug; methionine |
|
|
Term
- the genetic code is nearly __
- __ also use variations of the genetic code |
|
Definition
- universal
- mitochondria |
|
|
Term
-__ function as an adaptor molecule between a codon and an amino acid.
-There is at least one tRNA molecule for each __.
General characteristics of tRNA molecules include:
1.Each is a single strand of RNA between __ and __ amino acids in length.
2.The three-dimensional structure of the molecule is __.
3. In a two-dimensional representation, all tRNA molecules appear as a __ pattern. The amino acid–accepting region is the __, which contains the 3’ __ terminal region. Many of the nucleotides are involved in hydrogen bonds that form stems and loops.
4.Transfer RNA molecules contain unusual bases, such as __, or bases that have been modified.
5.The 5’ end is __ and the 5’ terminal residue is usually __.
6. The amino acid is attached to a __ of __ in the __ region of the acceptor stem.
7. The __ is in a loop near the center of the sequence. |
|
Definition
- tRNA
- amino acid
1. 73; 93
2. L-shaped
3. cloverleaf; acceptor stem; cca
4. inosine
5. phosphorylated; pG
6. hydroxyl group; adenosine; cca
7. anticodon |
|
|
Term
-Some transfer RNAs recognize more than one codon due to __ in base pairing
1.Codons that differ in either of the __ must be recognized by different tRNA.
2.The first base of the
anticodon the degree
of wobble. If the first
base is __, the
anticodon can recognize three codons |
|
Definition
- wobble
1. first two nucleotides
2. inosine |
|
|
Term
| synthesis of long proteins requires a low __ |
|
Definition
|
|
Term
- amino acids are activated by attachment to __
-To be incorporated into proteins, amino acids must be activated.
-Amino acids are activated by formation of an __ linkage between the carboxyl group of the amino acid and either the 2’ or 3’ hydroxyl group of the terminal adenosine of the tRNA, forming an aminoacyl tRNA or __ |
|
Definition
- tRNA
-
- ester; charged tRNA |
|
|
Term
-___ amino acids are covalently attached to the 3’ end of each tRNA molecule (named as: alanyl-tRNAAla)
-__ catalyze these reactions
-Most species have at least __ different aminoacyl-tRNA synthetases (1 per amino acid)
-__ are high-energy molecules (the amino acid has been “activated”)
-The activation of an amino acid by aminoacyl-tRNA synthetase requires __, both high energy phosphate bonds are broken |
|
Definition
- aminoacyl tRNA
- aminoacyl tRNA synthetases
- 20
- aminoacyl tRNAs
- atp |
|
|
Term
- amino acids are first activated by __
- the first step is the formation of __ or __
- the aminoacyl group is then transferred to a specific tRNA recognized by the __
- the aminoacyl amp never leaves the active site of the __ |
|
Definition
- adenylation
- aminoacyl adenylate; aminoacyl amp
- synthetase
- synthetase |
|
|
Term
-Attachment of the correct amino acid to the corresponding tRNA is a critical step
-__ binds ATP and the correct amino acid (based on size, charge, hydrophobicity)
-__ then selectively binds specific tRNA molecule based on structural features |
|
Definition
-
- synthetase
- synthetase |
|
|
Term
| - some aa-tRNA synthetases can be __ |
|
Definition
|
|
Term
-Synthetases are the true translators of the genetic code in that they assign a particular __ to a specific __.
-Many aspects of the tRNA molecule, in addition to the anticodon, are used as recognition sites by the __ to achieve this specificity. |
|
Definition
- amino acid; tRNA
- synthetases |
|
|
Term
-The __ is the site of protein synthesis.
-In E. coli, the ribosome sediments at 70S and is made of two subunits, the large __ subunit and small __ subunit.
-The 50S subunit = __
-The 30S subunit = __. |
|
Definition
- ribosome
- 50s; 30s
- 34 proteins + 23s RNA+ 5S RNA
- 21 proteins + 16S RNA |
|
|
Term
-Two-thirds of the mass of ribosomes is __, which is critical for the structure and function of the ribosome.
-The ribosomal RNAs fold into complex structures with many short duplex regions.
-Ribosomal RNA is the actual catalyst for __, with the ribosomal proteins making only a minor contribution. |
|
Definition
- RNA
-
- protein synthesis |
|
|
Term
-Because transcription and translation both occur in the __ direction, bacterial protein synthesis begins before transcription is complete.
-Several ribosomes can translate an mRNA molecule at the same time, forming __ or __ |
|
Definition
- 5>3
- polyribosomes; polysomes |
|
|
Term
- protein synthesis decodes the info in __
- protein synthesis is very ___
- protein synthesis consists of three parts: __, __, and __
-Initiation requires the cooperation of __, __, __, and proteins called __. – very complicated. |
|
Definition
- mRNA
- energetically costly
- intiation, elongation, and termination
- ribosomes, tRNA, mRNA, initiation factors |
|
|
Term
- Ribosomes have three tRNA binding sites
1.The __ binds the incoming tRNA.
2.The __ binds the tRNA with the growing peptide chain.
3.The __ binds the uncharged tRNA before it leaves the ribosome. |
|
Definition
1. A (aminoacyl) site
2. P (peptidyl) site
3. E (exit) site |
|
|
Term
-The __ holds the mRNA
-Each tRNA is in contact with both the __ and __ ribosomal subunits.
-The acceptor end of the tRNAs
in the __ and __ are near
one another – this is where the
__ is formed. A
channel connects this site to
the back of the ribosome
through which the peptide exits the ribosome during synthesis. |
|
Definition
- 30S ribosomal subunit
- 30S; 50S
- A site; P site; peptide bond |
|
|
Term
-In bacteria, the start signal is __ preceded by several bases that pair with __
-Many mRNAs in bacteria are __ - a single mRNA encodes for multiple proteins. Each of the coding regions has its own __.
-The first codon to be translated is usually __
-Initiation in bacteria begins at least 25 nucleotides from the __ end of the mRNA. |
|
Definition
- AUG (or GUG); 16S ribosomal RNA
-polycistronic; initiation site
- AUG (methionine)
- 5' |
|
|
Term
-The nucleotides between the 5’ end of the mRNA and the first codon translated are called the __
-The UTR contains a __ that directs protein synthesis machinery to the start site. The previous sequence is __, about 10 bp upstream of the start site, and interacts with the
__ to correctly position
the AUG codon in
the __. |
|
Definition
- untranslated region (UTR)
- shine dalgarno; purine rich; 16S rRNA; A site |
|
|
Term
- bacterial protein synthesis is initiated by __
-A modified form of methionine, __, is the initiator amino acid in most proteins in bacteria.
-It is attached to the __. __ binds only to the initiation codon (AUG- not to other AUGs.
-Another tRNA—__—recognizes internal codons for methionine.
-The same __ activates both tRNAm and tRNAf and a specific __ modifies the methionine attached to tRNAf |
|
Definition
- formylmethionyl tRNA
- n-formylmethionine (fMET)
- initiatior tRNA (tRNAf); fmet-tRNAf
- tRNAm
- synthetase; transformylase |
|
|
Term
-f-Met-tRNAf is placed in the __ during formation of the __ initiation complex
-__ assist in the assembly of the protein-synthesizing machinery.
-__ and __ bind the 30S subunit, preventing premature binding to __.
-__ brings fMet-tRNAf to the mRNA, forming the 30S initiation complex.
-The __ subunit binds, GTP is hydrolyzed by __, the initiation factors leave, resulting in the __ initiation complex. The fMet-tRNAf occupies the P site bound to AUG thereby establishing the reading frame. |
|
Definition
- ribosomal P site; 70S
- initiation factors (IF)
-IF1; IF3; 50S
- IF2 (with GTP)
- 50S; IF2; 70S |
|
|
Term
- elongation factors deliver __ to the ribosome
-__ is one of the most abundant proteins in bacteria
-__ inserted into __ site by __ during chain elongation
-Hydrolysis of GTP causes conformational change in __ – releases __ |
|
Definition
- aminoacyl tRNA
- EF-Tu
- aa-tRNA; A; EF-Tu
- EF-Tu; aa-tRNA |
|
|
Term
-Only __ form of EF-Tu can bind aa-tRNA.
-__ helps
__
Dissociate
-So one __ hydrolyzed for each tRNA brought by EF-Tu |
|
Definition
- GTP bound
- EF-Ts; GDP
- GTP |
|
|
Term
-The __ is now primed to catalyze peptide bond formation.
-Peptide bond formation, a thermodynamically favorable processs, is catalyzed by a site on the __ RNA, called the __. The 23S RNA is a component of the 50S subunit.
- __ catalyzes peptide bond synthesis |
|
Definition
- ribosome
- 23S RNA; peptidyl transferase center
- peptidyl transferase |
|
|
Term
| -Formation of a peptide bond – __ of amino group acts as nucleophile attacking __ of peptidyl-tRNA |
|
Definition
| - nitrogen; carbonyl carbon |
|
|
Term
-Formation of the peptide bond is followed by GTP-driven translocation of __ and __
- Elongation factor __ uses the energy of GTP hydrolysis to translocate the __ by one codon.
-Upon translocation, the peptidyl-tRNA is fully in the __, the __ is vacant, and uncharged tRNA is in the __, disengaged from the mRNA.
-The polypeptide chain grows from the __ terminal to the __ terminal. |
|
Definition
- tRNAs; mRNA
- G; mRNA
- P site; A site; E site
- amino; carboxyl |
|
|
Term
-Elongation continues with the growing chain exiting through a channel in the 50S subunit until a s__ appears in the __.
-Stop codons are recognized by __. __ and __ catalyze dissociation of the ribosome, mRNA, and attached tRNA in a reaction facilitated by __. |
|
Definition
- stop codon; a site
- release factors (RF); EF-G; ribosome release factor (RRF); gtp hydrolysis |
|
|
Term
| - protein synthesis is energetically expensive, __ are cleaved for each amino acid added to a polypeptide chain |
|
Definition
| - four phosphoanhydride bonds |
|
|
Term
Eukaryotic protein initiation is different
1.The ribosomes are larger - __ and __ subunits that form the __ ribosome.
2.Protein synthesis begins with a __ rather than __. A special initiator tRNA called __ is required.
3.The initiator codon is always the first __ from the 5’ end of the mRNA. More protein __ are required.
4. the mRNA is __ because of interactions between proteins that bind the 5' cap and those that bind the poly a tail
5.Elongation and termination are similar except that bacteria have __ release factors while eukaryotes have only __.
6.Eukaryotic protein synthesis occurs in the __ and utilizes large complexes associated with the __. |
|
Definition
1. 40S; 60S; 80S
2. methionine; formylmethionine; met-tRNAi
3. aug; initiation factors
4. circular
5. two; one
6. cytoplasm; cytoskeleton |
|
|
Term
| Mutations in eukaryotic initiation factor 2 result in __ disease, characterized by the disappearance of brain nerve cells which are replaced by cerebrospinal fluid. |
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Definition
| vanishing white matter disease |
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Term
-Many antibiotics inhibit __ while leaving __ unaffected.
-___ interferes with the binding of fMet-tRNAf and thereby inhibits protein synthesis initiation in bacteria |
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Definition
- bacterial protein synthesis; eukaryotic protein synthesis
- streptomycin |
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Term
- some antibiotics inhibit protein synthesis
--__ resembles the 3’ end of an aminoacyl-tRNA, and can enter the A site of a ribosome - The peptidyl-__ formed is bound weakly in the A site and dissociates terminating protein synthesis |
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Definition
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Term
Diptheria toxin blocks protein synthesis in eukaryotes by inhibiting __
-Corynebacterium diphtheriae, which causes diphtheria, grows in the upper respiratory tract of an infected individual and produces a toxin that inhibits __.
-The toxin covalently attaches ADP-ribose to an amino acid in elongation factor 2, preventing __ and consequently protein synthesis.
-In unimmunized individuals, infection can be fatal. |
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Definition
translocation
- protein synthesis
- elongation
- |
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Term
-__ is a small highly toxic protein found in castor beans.
-Ricin has a catalytic activity that cleaves __ from a nucleotide in __ that is crucial for binding elongation factors. Protein synthesis is halted. |
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Definition
- ricin
- adenine; 28s RNA |
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Term
-In eukaryotes, protein sorting directs proteins to distinct organelles.
-In the secretory pathway, proteins are inserted into the __ membrane co-translationally.
-Protein synthesis in the secretory pathway occurs on ribosomes bound to the __. ER with ribosomes bound is called the __. |
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Definition
-
- er membrane
- er; rough er (rer) |
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Term
-The synthesis of proteins bound for the secretory pathway begins on __ that are free in the __.
-First, a portion of the nascent protein that contains a specific signal (__) emerges from the ribosome. The signal peptide is a sequence of 9 to 12 __, often located at the __ region of the primary structure
-The __ binds the signal sequence as it exists the ribosome and directs the complex to the ER. Binding of __ to the ribosome halts protein synthesis. |
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Definition
- ribosomes; cytoplasm
- (the signal peptide); hydrophobic amino acids; n-terminal
- signal recognition particle (srp); srp |
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Term
-Next, the __, an integral membrane with __, binds to the SRP-ribosome complex.
-The __, a protein-conducting channel, accepts the ribosome from the SRP-SRP receptor complex and __ begins again with the protein now passing through the membrane in the translocon |
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Definition
- srp receptor; GTPase
- translocon; protein synthesis |
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Term
mRNA use is subject to regulation
-Iron transport and storage is carefully regulated.
-__: an iron storage protein in the cell, has translational regulation from its mRNA. A stem-loop structure in the 5’ UTR (iron response element or IRE) binds the IRE-binding protein (IRE-BP) when iron is absent and prevents translation. When present, iron binds the IRE-BP causing it to dissociate from the IRE, allowing translation occur. |
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Definition
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Term
-__, which binds iron-rich transferrin and facilitates its entry into cells, also several IREs located in the 3’ untranslated region.
-When iron is absent, __ binds the __, allowing the mRNA to be translated.
-When iron is present, it binds to the __, which dissociates from the transferrin-receptor mRNA. Without the __, the receptor mRNA is degraded |
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Definition
- transferrin-receptor
- ire-bp; IREs
- IRE-BP; IRE-BP |
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Term
-__ leads to mRNA degradation induced by the presence of double-stranded RNA, which may be present during certain viral infections.
-__, a ribonuclease, cleaves double-stranded RNA into small fragments, called __ that are bound by a class of proteins called __ to form the RNA induced silencing complex (RISC).
-The complex locates mRNA complementary to the __ and degrades the mRNA. |
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Definition
- RNA interference (RNAi)
- dicer; small interfering RNA (siRNA); argonaute
- siRNA |
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Term
MicroRNA
-MicroRNAs (miRNAS) – __ long
-Regulate gene expression __
-May regulate as much as __ of mammalian protein-coding genes
-Regulate mRNA __ and/or __
-Some miRNAs promote __ of specific mRNAs
-Some miRNAs promote __ or __ of mRNAs |
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Definition
- 21 nucleotides
- post-transcriptionally
- 60%
- stability; translation
- cleavage
- translational repression; deadenylation |
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Term
-__ can be used to cleave DNA is specific places.
-These bacterial enzymes recognize cleavage sites of four to eight nucleotides in length that are __, then cleave each strand of the DNA. |
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Definition
- restriction enzymes
- palindromic |
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Term
-DNA fragments generated by restriction enzyme digestion can be separated by gel electrophoresis in __ or __
-The fragments can be visualized by staining with __.
-Specific fragments can be isolated from the gel. |
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Definition
- agarose; polyacrylamide gels
- ethidium bromide |
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Term
- __, in which all but one reactive group is blocked, can be sequentially linked to a strand attached to an insoluble support, making DNA artificially!
-The newly synthesized DNA can be tagged with 32P or a fluorescent marker
and used as a __,
or could be used as
a __ |
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Definition
-Deoxyribonucleoside3’-phosphoramidites
- probe; primer |
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Term
-To identify a specific sequence, the separated fragments are transferred to nitrocellulose paper, a process called __, and then exposed to a radioactively labeled probe complementary to the sequence of interest.
-__ identifies the fragment which binds the probe |
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Definition
- southern blotting
- autoradiography |
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Term
-__ and __ are key tools for forming recombinant DNA molecules
-Digestion of DNA by __ often generates staggered or sticky ends.
-A __ (a piece of DNA readily taken up and replicated by bacteria) is cleaved by the same restriction enzyme. The ends of the fragment and cleaved vector can anneal, and __
is used to covalently join
them, generating
recombinant DNA |
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Definition
- restriction enzymes; dna ligase
- restriction enzymes
- vector; dna ligase |
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Term
-__ are small, circular DNA molecules used as vectors for DNA fragments to 20kb
-__ autonomously within a host cell
-Carry genes conferring __, used as marker genes for cells carrying vectors
-Generally the best choice for small inserts
-pBR322 was one of the first plasmid vectors - many of the vectors used today were derived from pBR322 |
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Definition
- plasmids
- replicate
- antibiotic resistance |
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Term
- __: Efficient, commonly used vector for delivering DNA into a bacterial cell - especially important for screening libraries - clone insert into a bacteriophage
-Advantage over plasmid vectors is that __ is more efficient than __ - especially for large DNA fragments
-Disadvantage: DNA must be packaged into phage particles __ - can’t have a line of cells that propagates the molecule |
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Definition
- bacteriophage lambda vectors
- transfection; transformation
- in vitro |
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Term
-__ can replicate in either prokaryotic or eukaryotic cells
-They can be used to transfer __ between prokaryotes and eukaryotes
-Useful for cloning __ in bacteria (can easily manipulate DNA here), and then expressing the gene products in a eukaryotic cell (many eukaryotic proteins are not properly modified in bacterial cells) |
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Definition
- shuttle vectors
- recombinant dna
- eukaryotic dna |
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Term
-After a cloning vector and insert DNA have been joined in vitro, __ is introduced into a host cell such as E. coli (transformation)
-Only a small percentage of cells take up the DNA
-__: cells are grown under conditions in which only transformed cells survive
- __: transformed cells are tested for presence of the recombinant dna |
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Definition
- recombinant dna
-
- selection
- screening |
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Term
six steps in a recombinant dna experiment
1. Preparation of DNA. __ and __
2. Cleavage of __ at __. Insert DNA can be added at specific points that have been cleaved
3. __ of DNA fragments. Joining of the fragments
4. Introduction of __ into compatible host cells. (Genetic transformation)
5. __ and __ of recombinant DNA in host cells. __ allow insert DNA to be replicated in host cells
6. __ of host cells that contain recombinant DNA of interest. Screening a large number of DNA clones for desired fragment |
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Definition
1. dna vector; target dna
2. dna; particular sequences
3. litigation
4. recombinant dna
5. replication; expression; cloning vectors
6. identification |
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Term
-__ is a double-stranded DNA molecule generated from an mRNA template by __, a retroviral enzyme.
-mRNA is specifically targeted via its __. |
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Definition
- complementary DNA (cDNA); reverse transcriptase
- poly A tail |
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Term
-__ can be synthesized from all of the mRNA that a cell contains and inserted into a cloning vector to generate a __.
-Clones containing __ from eukaryotes can be expressed in bacteria.
-__ represent all the mRNAs made in a given cell or tissue – so they vary depending on cell type they were constructed from.
-cDNA libraries from mRNA do not include __ or __ (much less complex than genomic libraries) |
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Definition
- complementary DNA; cDNA library
- cDNA
- cDNA libraries
- introns; flanking sequences |
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Term
-__ complementary to the desired sequence can be used to screen a cDNA library.
-__ containing a cDNA library are used to infect bacteria growing on a petri plate. Sites of infection, called __, contain identical phages.
-A replica of the petri plate is made on nitrocellulose, the DNA is denatured with NaOH and the __ is used to identify plaques containing the cDNA of interest |
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Definition
- probes
- lambda phage; plaques
- radioactive probe |
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Term
-Complementary DNA can also be inserted into __, which have powerful promoters for transcription as well as the signals needed for translation
-Replica plates are probed with an __ for the protein of interest. |
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Definition
- expression vectors
- antibody |
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Term
-A __ is made by fragmenting the entire genome of an organism and inserting the fragments into a vector such as λ phage.
-Bacteria are infected with the λ phage, replicas appear and are then screened using radioactive probes for the gene of interest.
-Genomic libraries do not vary between __, and contain __ and __ |
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Definition
- genomic library
-
- tissues; introns; non-coding dna |
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Term
-__: plasmids that have been engineered to contain regulatory sequences for transcription and translation (strong promoters, ribosome-binding sites, transcription terminators)
-Eukaryotic genes can be expressed in prokaryotes - get expression, though modifications not the same. |
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Definition
-prokaryotic expression vectors
- |
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Term
expression of proteins in eukaryotes
-Prokaryotic cells may be unable to produce functional eukaryotic proteins, __ are not the same.
-Some __ are for eukaryotes - contain eukaryotic origins of replication, marker genes, etc.
-These can be transfected into eukaryotic tissue culture cells and the expressed proteins studied or purified.
-Recombinant viruses (adenovirus, vaccinia virus) can also be employed to express proteins in eukaryotic cells |
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Definition
- post translational modifications
- expression vectors
-
- |
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Term
-__ molecules can also be integrated into the genomes of large multicellular organisms
-Creates __: organisms with stably integrated foreign DNA |
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Definition
- recombinant dna
- transgenic organisms |
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Term
-DNA can be sequenced by controlled termination of __
-__ uses 2’,3’-dideoxynucleoside triphosphates (ddNTPs) which are incorporated at the 3’ end of a growing chain in place of a dNTP
-Since ddNTPs lack a __
group, subsequent nucleotide
addition cannot take place
-Small amounts of ddNTP’s terminate replication of some chains at each step, leaving a set of fragments of different lengths
-Results can be read on a gel (if radioactive sample used), or on a fluorimeter (if fluorescently labeled dNTPS are used) |
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Definition
- replication
- sanger method
- 3' hydroxyl |
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Term
-The location to be sequenced is determined by the added __
-Strands of different lengths will result, with the last nucleotide added always the dideoxy analog.
-Separation on a gel that allows detection of one nucleotide differences in size allows you to read the sequence |
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Definition
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Term
-The __ is used for amplifying a small amount of DNA - can get enough to study from a very small sample
-Also can increase the proportion of a particular DNA sequence in a mixed DNA population
-Used is analysis of medical samples: diagnosis of infectious diseases, determination of genetic defects, paternity questions, matching of donor organs
-Also used in study of molecular evolution |
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Definition
| - polymerase chain reaction (PCR) |
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Term
-PCR is capable of __ a single molecule of DNA which can then be visualized in a gel or sequenced.
-PCR is used to detect __ in genes that cause cancer, verify the presence of infectious agents in cells, and determine the guilt or innocence of crime suspects. |
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Definition
|
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Term
site directed mutagenesis of cloned dna
-Powerful technique to introduce a desired __ directly into a gene
-__ is synthesized containing mutation and flanking sequences of gene
-From __, DNA replication produces a new copy of the mutated gene
-Important in structure-function studies of genes and their protein products |
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Definition
- mutation
- oligonucleotide
- oligonucleotide primer |
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Term
-The quantity of individual transcripts in a cell can be determined by __
-__ (made from mRNA) is used in PCR reaction along with a dye that fluoresces when binding duplex DNA. The PCR cycle at which fluorescence's becomes detectable is inversely related to the original number of target templates. |
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Definition
- quantitative PCR (qPCR, real-time PCR)
- cDNA |
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Term
| __ or gene chips allow the determination of the expression pattern of a large number of genes simultaneously. cDNAs are attached to a solid support in a defined pattern to generate the microarray. Fluorescently labeled cDNA is then hybridized to the microarray to reveal the expression level of each gene on the chip. |
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Definition
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Term
steps for PCR:
1. Separate DNA duplex by heating to __
2. Add __ to desired region (must know sequence for primers to anneal to!)
3. Add __ (Taq, Vent) - allow replication for a short period.
-__ strands again; repeat __ and __ steps |
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Definition
1. 95C
2. primers
3. dna polymerase
- heat; annealing; replication |
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