Term
| Which of the three models of DNA replication proposed that the original double helix remained entirely intact throughout the process? |
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Definition
-CONSERVATIVE: eliminated after 1st round
(semiconservative was correct model) |
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Term
DNA replication begins?
-and travels? |
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Definition
| begins at origin & travels bidirectionally |
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Term
| Why is an AT rich region part of the E. coli oriC? |
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Definition
| AT base pairs (2 H bonds) are more easily separated than GC base pairs (3 H bonds). |
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Term
| acts as template strand in DNA replication? |
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Definition
| the original DNA strand aka parental |
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Term
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Definition
| nucleotides (4 bases: A, T, G, C) |
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Term
| responsible for holding double stranded DNA together? |
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Definition
-base stacking
-H bonding between opposite strands (AT & GC) |
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Term
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Definition
antiparallel
-which is determined by orientation of sugar molecules in sugar-phosphate backbone of DNA
-one strand is 5'->3'; other strand is 3'->5' |
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Term
| H bonding between nucleotides and template strands follow? |
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Definition
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Term
| ______ strand travels toward replication fork |
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Definition
leading strand
-synthesized continuously |
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Term
| _______ strand travels away from replication fork |
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Definition
lagging strand
-made of Okazaki fragments |
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Term
| bacterial DNA has how many replication forks? |
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Definition
| two (& 1 origin of replication) |
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Term
oriC
-means?
-3 types of DNA sequences within? |
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Definition
-origin for chromosomal replication
1. AT rich region
2. DnaA box sequences (5)
3. GATC methylation sites |
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Term
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Definition
| there are 3 tandem repeats that are 13 bp long |
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Term
| method by with DNA replication was shown to be semiconservative? |
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Definition
| -by labeling DNA with heavy and light isotopes of nitrogen and using centrifugation |
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Term
| In E. coli, DNA replication is initiated when? |
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Definition
-DnaA proteins bind to 5 DnaA boxes at origin and cause AT rich region to unwind.
-DNA helicase (recruited by DnaA and DnaC proteins) then breaks H bonds between strands & promote movement of two replication forks. |
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Term
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Definition
|
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Term
| alleviates supercoiling caused by DNA helicase? |
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Definition
topoisomerase (type II) aka DNA gyrase
-travels in front of DNA helicase |
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Term
| Parental strands remain separated via? |
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Definition
single-stranded proteins
-prevent reforming of dsDNA by coating ssDNA |
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Term
RNA primers
-definition?
-by what? |
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Definition
=short strands (10-12 nucleotides) of RNA synthesized by linkage of ribonucleotides
-PRIMASE
single primer @ origin in leading strand; multiple primers in lagging strand |
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Term
| synthesizes DNA of leading & lagging strands (aka daughter strand) in bacterial DNA replication? |
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Definition
DNA polymerase (I & III)
-mostly III (10 subunits)
-I (1 subunit- removes RNA primers, fills with DNA) |
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Term
| removes RNA primers in bacterial DNA replication? |
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Definition
DNA polymerase I
-removes in 5'->3' direction
-fills vacant area using 3' end of Okazaki fragments |
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Term
| Which enzyme produces the first piece of nucleic acid synthesized in E. coli DNA replication? |
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Definition
| DNA Primase (synthesizes RNA primers) |
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Term
| first piece of nucleic acid? |
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Definition
RNA primers
-DNA polymerase need a primer to synthesize DNA & make new DNA strands in 5'->3' direction |
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Term
seals the gaps between Okazaki fragments in bacterial DNA replication?
-type of energy? |
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Definition
DNA ligase
-requires NAD+ in E. coli
-requires ATP in archaea and eukaryotes |
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Term
| DNA gyrase function in bacterial DNA replication? |
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Definition
aka topoisomerase (type II)
alleviates positive supercoiling caused by DNA helicase dsDNA separation |
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Term
| alpha subunit of DNA polymerase III function? |
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Definition
| -catalyze bond formation between adjacent nucleotides |
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Term
| all 10 subunits of DNA polymerase III aka? |
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Definition
| DNA polymerase III holoenzyme |
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Term
| enzyme that wraps around DNA like a hand? |
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Definition
DNA polymerase III
-DNA threaded through palm
-dNTPs enter catalytic site, bind to template (AT/GC rule) & are covantely attached to 3' end of growing strand |
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Term
| DNA polymerases synthesize DNA in which direction? |
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Definition
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Term
processive enzyme
-definition
-example |
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Definition
=enzyme, such as RNA or DNA polymerase, which glides along the DNA & does not dissociate from the template strand as it catalyzes the covalent attachment of nucleotides
ex) DNA polymerase III (uses dNTPS to make new DNA strands) |
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Term
| 3' exonuclease site found in? |
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Definition
DNA polymerase III
-removes mismatched bases |
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Term
| DNA polymerase III catalyzes attachment of nucleotides to ___ end of each primer? |
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Definition
| 3' (in 5'-> 3' direction) |
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Term
| dNTPs attached to __ end of growing strand in DNA polymerase III? |
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Definition
| 3' (in 5'-> 3' direction) |
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Term
| RNA primer at ___ end of DNA in lagging strand? |
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Definition
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Term
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Definition
Okazaki fragments
-covalent bond between last nucleotide added by polymerase I & adjacent DNA strand previously made by polymerase III |
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Term
| DNA polIII requires a ________ in order to continue elongation of the strand |
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Definition
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Term
| What is a key difference between DNA polIII and DNA ligase? |
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Definition
| DNA ligase can use energy from ATP rather than nucleotides. |
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Term
| explanation for Okazaki fragments |
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Definition
| DNA polymerase requires a free 3' OH group to attach new nucleotides; primers are produced on the lagging strand every 1000-2000 nucleotides. |
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Term
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Definition
3 phosphate groups attached to 5' carbon atom of deoxyribose
-breaks covalent bond betwen 2 phosphates (highly exergonic)
-->provides energy to form covalent (ester) bond between sugar at 3' end of DNA & phosphate of incoming nucleotide
--->strand grows 5->3 & PPi release |
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Term
phosphodiester linkage
-aka?
-means?
-describes? |
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Definition
aka phosphodiester bond
-means 2 ester bonds
-describes linkage between phosphate and 2 sugar molecules |
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Term
| protein, Tus, binds to ______ & stops movement of replication forks |
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Definition
| Tus (termination utilization substance) binds to ter (termination) sequences |
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Term
| What DNA sequence is required for termination of replication in E. coli? |
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Definition
ter sequences (termination sequences)
-located on opposite side of oriC
-Tus=protein that binds there |
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Term
| What is the major difference between T1 and T2 ter sequences? |
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Definition
| T1 stops only counterclockwise-moving forks while T2 stops only clockwise-moving forks. |
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Term
| T1 stops ______-moving forks |
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Definition
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Term
DNA replication can sometimes lead to two intertwined DNA molecules known as?
-separated by? |
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Definition
-catenanes (transient structures)
-topoisomerase II introduces a temporary break into DNA strands & rejoins after unlocked |
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Term
| type of bond that occurs between adjacent nucleotides? |
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Definition
single covalent ester bond
(vs. phosphodiester between phos & 2 sugars) |
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Term
| DNA polymerase III alpha function? |
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Definition
|
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Term
| DNA polymerase III epsilon function? |
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Definition
| 3'->5' proofreading that removes mismatched nucleotides |
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Term
| DNA polymerase III theta function? |
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Definition
| accessory protein that stimulates the proofreading function (epsilon) |
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Term
| DNA polymerase III beta function? |
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Definition
clamp protein
-allows DNA polymerase to slide along the DNA without falling off |
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Term
| DNA polymerase III T, y, delta, delta', psi, chi function? |
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Definition
clamp loader complex
-helps clamp protein (beta) bind to DNA |
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Term
primosome
-composed of?
-importance? |
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Definition
-DNA helicase & primase bound to each to form complex
-complex leads the way at the replication fork
-->separates DNA & synthesizes RNA primers at regular intervals along lagging strand |
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Term
|
Definition
-primosome (helicase & primase) associate with 2 DNA polymerase holoenzymes
-can be found at the replication fork during DNA synthesis
-dimeric DNA polymerase (2 DNA polymerase holoenzymes that move as unit toward replication fork--> lagging strand must be looped out) --> hopping |
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Term
| how many ter sequences are necessary to stop advancement of replication fork in any cell? |
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Definition
only one
-DNA replication ends when opposite fork meets ceased fork. |
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Term
| proofreading function of DNA polymerase? |
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Definition
-the ability to remove mismatched bases in 3'->5' direction at the 3' exonuclease site
AKA exonuclease cleavage |
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Term
| mechanisms by which E. coli can coordinate DNA replication with cell division? |
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Definition
1. DnaA protein does not efficiently bind to hemimethylated binding sites in the oriC.
2. DnaA protein is rapidly degraded after the initiation of DNA replication.
3. DnaA proteins must bind to a minimum number of binding sites in the oriC. |
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Term
| DnaA protein is unable to bind hemimethylated DNA because its binding site includes a GATC methylation site? T/F |
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Definition
| False, While there are many GATC methylation sites in the oriC, and full methylation of these sites is required for efficient binding of DnaA protein, the DnaA box does not actually include GATC methylation sites. |
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Term
| The fidelity of DNA replication in E. coli is about one mistake per ___________ incorporated nucleotides |
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Definition
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Term
| DNA polymerase uses energy contained in the linkage between phosphate groups to drive DNA synthesis? T/F |
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Definition
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Term
| After DNA replication, the newly synthesized strand contains only newly synthesized histones? T/F |
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Definition
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Term
| You extract DNA from an E. coli cell and observe it is hemi-methylated. Which strand of DNA is older? |
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Definition
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Term
| What types of mutants were essential to the discovery of new replication enzymes? |
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Definition
| Temperature-sensitive mutations (which is a type of conditional mutant) |
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Term
| Lesion-replicating polymerases are _________. |
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Definition
| Used to replicate damaged DNA in Eukaryotes |
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Term
| The problem is synthesizing the 3' end of chromosomes is solved by the use of ?? |
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Definition
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Term
| What covalently attaches a methyl group to adenine to form methyladenine? |
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Definition
| Dam (DNA adenine methyltransferase) |
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Term
Prior to DNA vs. after DNA
-what is methylated? |
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Definition
Prior: Dam methylates both adenines within GATC sites
After: only adenines in original strands are methylated. Will take several minutes until Dam methylates the unmethylated adenines |
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Term
|
Definition
autonomously replacting sequence
-origins in eukaryotes, S. cerevisiae
-have high % of AT bases, copy of ARS consensus sequence (ACS)
-->enhance origin function |
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Term
| yeast origin determined by? |
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Definition
|
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Term
| animal orgins determined by? |
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Definition
| specific sites along a choromose due to chromatin structure & protein modifications |
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Term
| Bacterial DNA replication regulated by: |
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Definition
1. amt of DnaA protein
2. methylation of GATC sites in oriC |
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Term
| high fidelity of DNA is result of? |
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Definition
fidelity=accuracy
1.stability of H bonds btwn correct bases
2. phenomenon of induced fit
3. proofreading ability of DNA polymerase |
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Term
| DNA replication licensing |
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Definition
-when MCM helicase binds at origin
--> initiates DNA synthesis in Eukaryotes |
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Term
| DNA replication in Eukaryototes begins with? |
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Definition
assembly of preRC (prereplication comples)
-14 proteins
-part of preRC: ORC (origin recognition complex)- 6 proteinds
-->ORC binds to ARS |
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Term
| Euk DNA replication is usually initiated when? |
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Definition
-G1 phase
-when ORC of preRC binds to ARS |
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Term
| Euk DNA synthesis begins when? |
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Definition
-During S phase
-when preRC is acted upon by 22 proteins -->activates MCM helicase & assemble 2 divergent forks |
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Term
| DNA polymerases involved with Eukaryotic DNA replication in cell nucleus during S phase? |
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Definition
|
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Term
| DNA polymerase that functions with mitochondria to replicate mitochondrial DNA? |
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Definition
|
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Term
|
Definition
-associates with primase
-->DNA poly a/primase complex synthesizes short RNA-DNA primer.
--->then dissociates from fork and is exchanged for epsilon or delta poly |
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Term
| polymerase switch during eukaryotic DNA replication? |
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Definition
After DNA poly a/primase complex synthesizes RNA-DNA primers, dissociates from fork. Exchanged for epsilon (leading strand) or delta (lagging strand)
-during S phase |
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Term
| DNA poly beta in euk DNA rep? |
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Definition
| removes incorrect bases from damaged DNA |
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Term
| RNA primers in Euk DNA replication removed by |
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Definition
flap endonuclease
-removes flaps from RNA primers on Okazaki fragments |
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Term
|
Definition
| elongates Okazaki fragments until it runs into RNA primer of adjacent Okazaki |
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Term
|
Definition
cleaved by Dna2 nuclease/helicase
-then removed by flap endonuclease |
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Term
|
Definition
| =comples of telomeric sequences (moderately repetitive tandems & a 3' overhang region) at both ends of linear D˜Å |
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Term
|
Definition
=prevents chromosomal shortening using a short RNA molecule as a template to add repeat sequences onto telomeres
-binds to 3' overhang |
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Term
| After strand is synthesizes in telomeric region, what synthesizes complementary strand? |
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Definition
| primase, DNA polymerases, & DNA ligase |
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