Entry of virus nucleocapsid into host cytoplasm

Reverse Transcription of viral RNA into DNA by viral reverse transcriptase

Integration of viral DNA with host DNA genome = provirus

Transcription of proviral DNA by host

Polymerase II in host nucleus and subsequest transportation to cytoplasm for translation

Assembly, Budding & Maturation of HIV into infectious particles

Which region in the HIV genome codes for protease, reverse transcriptase, ribonuclease, and integrase?

Which region codes for nucleocapsid core and matrix proteins p24, p17, p7 and p6?

viral enxymes:  The POL region

structural proteins: The GAG region

NEF

-encodes negative regulatory factor

-deletion of non-essential NEF gene represents strategy for possible live attenuated HIV vaccine

What type(s) of eukaryotic RNA polymerase are inhibited by a-amanitin, a toxin found in the poisonous mushroom Amanita phalloides? Where is each polymerase located?

A. RNA pol I

B. RNA pol II

C. RNA pol III

A. RNA pol I - Nucleolus

B. RNA pol II - Nucleoplasm

C. RNA pol III - Nucleoplasm

RNA polymerases II and III are inhibited by a-amanitin - II is strongly inhibited, III in high concentrations, and I not at all.

CBP

(CREB binding protein)

2 meters

23 pairs of chromosomes

N-linked oligosaccharides, characteristic of plasma and membrane proteins, are always attached by a glycosylamine linkage of N-acetylglucosamine to what?

What channels open during repolarization?

Voltage-gated Na+ channels

ex. lidocaine used as local anaesthetic - inhibits Na+ channel opening and depolarization of cell

- cocaine also inhibits dopamine and norEpi reuptake

What are the 5 classes of Neurotransmitters?

Examples of each?

Amines: ACh, dopamine, serotonin

Amino Acids: Glutamate, glycine, GABA

Purines: ATP, adenosine

Gases: NO

Peptides: opioid peptides, somatostatin

How is ACh made?

How is it broken down?

Acetyl group is combined with choline by  choline acetyltransferase --> ACh

ACh is broken down into choline and acetate by acetylcholinesterase

What happens in Myasthenia Gravis?

- antibodies are made against nicotinic ACh receptors in muscle

- treated with drugs that inhibit acetylcholinesterase

- characterized by muscle weakness - ptosis

Cholera binds to the alpha subunit of the Gs protein and inhibits the intrinsic GTPase activity.

This causes constituitively active G protein --> elevated cAMP levels in intestinal epithelium and resulting diarrhea.

What is the rate-limiting step for catecholamine biosynthesis?

The conversion of tyrosine to DOPA by

Tyrosine Hydroxylase

- first step in the reaction

The inactivation of catecholamines by monoamine oxidase (MAO)

Dopamine --> DOPAL

Norepi & Epi --> DOPEGAL

substantia nigra

- etiology of degeneration unknown

What do the following genes all share in common?

DJ-1

a-synuclein

UCHL1

Parkin

PINK1

mutations in these genes causes rare inherited form of Parkinson's disease

-associated with oxidative stress & oxidative damage

What is the major excitatory neurotransmitter in the brain?

The major inhibitory one?

excitatory - glutamate

inhibitory - GABA

- GABA synthesized from glutamate via decarboxylation

Tryptophan

Trp  --->   5-Hydroxy-tryptophan --->  Serotonin

   hydroxylase                     Decarboxylase

b-secretase and g-secretase generate Ab peptide

- thought to cause Alzheimer Disease

- cleavage by a-secretase does not produce Ab peptide

XII ---> XIIa (cleaved by kininogen & kallikrein)

XI ---> XIa

IX ---> IXa

X ---> Xa

(note order: 12, 11, 9, 10)

What role does Vitamin K play in blood clotting?

What drug inhibits it?

Vitamin K allows Prothrombin ----> Thrombin

By adding g-carboxy group to a glutamate in Prothrombin,  Ca++ can bind.

Vit K also adds a g-carboxy group to factors IX, X, and VII. 

Coumadin (Warfarin, Dicoumarol) inhibits Vit K.

Why would heparin be given to a heart attack patient?

What type of heterpolysaccharide+protein molecule is heparin?

Heparin enhances Antithrombin III activity, preventing blood clot formation.

It is a glycosaminoglycan.

Serine Protease

- activates plasmin which lyses blood clot

Acetylation = decompacted = more transcription

deacetylation = tightly wound = less transcription

For transcription of nuclear DNA which nucleic acid sequence(s) correspond to "start"? To "stop"?

Which corresponds to "stop" for mtDNA?

Nuclear:            Start codon: AUG 

Stop codons: UAA, UGA, UAG

mtDNA:                Stop: AGA

What codon codes for methionine in nuclear DNA?

In mtDNA?

Nuclear DNA Met: AUG

mtDNA Met: AUA

A Silent Mutation

ex. if UCA (serine) was mutated to UCU (also serine)

- depends on the redundancy (degeneracy) of the code

- lengths of mutant proteins are unchanged

Which type of mRNA point mutation always results in a shorter mutant protein?

Which type always results in a longer protein?

Shorter:          Nonsense mutation 

- mutation results in termination codon

(UAA, UGA, UAG)

Longer:           Supressor mutation

- mutation results in suppression of termination

a Missense Mutation

GAG --> GUG

(Glu --> Val)

- affects the Hemoglobin Beta chain

What sized subunits are in Prokaryotic ribosomes?

Eukaryotic ribosomes?

Prokaryotic: 30, 50, 70

Eukaryotic: 40, 60, 80

True or False:

Eukaryotic mRNA is polycistronic, has a poly A tail, and a 7-methyl G noncoding cap.

False

- eukaryotic mRNA is monocistronic

the first reaction, catalyzed by 

5-aminolevulinate synthase:

Glycine + Succinyl CoA --> ALA + CO2 + CoA

- pyridoxal phosphate is required coenzyme

Ferrochelatase

- the last step, leading to the formation of heme, where Fe2+ is added to Protoporphyrin IX

ALA dehydrase

- the second step, resulting in formation of  porphobilinogen

5-aminolevulinate synthase

aka

ALA synthase

- this enzyme is catalyzes the first and rate-limitng reaction: formation of ALA from Succinyl CoA and Glycine in the mitochondria

deficiency of Uroporphyrinogen I synthetase

- AIP patients are not light-sensitive (does not affect skin)

- can result in urine that turns blue/black on standing

Decreased production of heme = increased ALA Synthase

(since hemin product is feedback inhibitor of this enzyme, its absence causes increased ALA synthase and enzyme activity, leading to buildup of intermediates)

What is the most common Porphyria?

What enzyme deficiency causes it?

Porphyria Cutanea Tarda

-lack of Uroporphyrinogen Decarboxylase

-patients are photosensitive

Hemin activates the synthesis of globin peptide

- increses globin while decreasing heme

What is Hemolytic (prehepatic) jaundice?

Hemolytic (prehepatic) jaundice: production of bilirubin exceeds liver's capacity to conjugate it w/ glucuronic acid for excretion --> unconjugated bilirubin is elevated in the blood.

Hemolytic anemia (caused by massive lysis of red blood cells, as in Sickle Cell), can lead to buildup of bilirubin, causing jaundice.

Bilirubin glucuronyl-transferase

-increases solubility of bilirubin

Heme (+ 02, NADPH + H+) -----------> Biliverdin

                     Heme oxygenase

Biliverdin (+ NADPH + H+) -----------> Bilirubin

                       Biliverdin reductase

decrease in blood pH (Bohr effect)

increase in blood levels of 2,3 BPG (high altitude)

True or False:

O2, CO2, CO, and glucose can all bind to hemoglobin

 True

O2: higher affinity when Hb in relaxed state

CO2: binds at tissues, transported as NHCOO-

CO: poison, 200x higher affinity than O2

& prevents O2 release

glucose: HbA1c levels diagnostic for diabetics

Hereditary defects in synthesis of either a or b chains

- common in Mediterraneans

- b defects more common in U.S.

Which usually contains more protein than carbohydrate: proteoglycans or glycoproteins?

Which contains more carbohydrate?

Glycoproteins: usually more protein

Proteoglycans: more carbohydrate

List 3 properties of Glycosaminoglycans (mucopolysaccharides).

1. N-containing

2. Polyanionic

(due to carboxyl-containing sugars [except Keratan sulfate] and/or sulfate groups)

3. Repeating disaccharide motif on long polysaccharide chain

Hyaluronic acid

Heparin

Heparin sulfate

Keratan sulfate

Chondroitin sulfate

Dermatan sulfate

glycosaminoglycans

(GAGS)

- all glycosaminoglycans except hyaluronic acid are found covalently attached to protein, forming a "bottle brush" proteoglycan monomer.

Osteogenesis Imperfecta (OI) aka "brittle bone disease"

- point mutation in COL1A1 or COL1A2 results in switching of Gly --> prevent folding to triple helix

- severity: II>III>IV>I

Ehlers-Danlos Syndrome (EDS)

- a collection of defects including skin and joint hypermobility, bruising, scoliosis

Which amino acids make up collagen's primary structure?

Gly-X-Y

Glycine at every 3rd position,

 X is often proline and

Y is usually hydroxyproline or hydroxylysine

What type of collagen is found in intervertebral discs, cartilage, and vitreous body?

Which type is found in fetal skin?

Which type is found in skin and bone, tendon, blood vessels, and cornea?

Type II - cartilage, etc.

Type III - fetal skin

Type I - skin and bone, etc.

(Type IV in basement membrane)

True or False:

Like collagen, elastin is rich in hydroxyproline and hydroxylysine residues.

False

Elastin contains proline and lysine, but little hydroxyproline and no hydroxylysine. Also rich in nonpolar aa's Gly, Ala, and Val.

In what fibrous protein are desmosine cross-links found?

What are they?

What do they do for the macro-stucture of the protein?

-Desmosine cross-links found in elastin.

-Consist of 3 allysine chains + 1 lysyl side chain

-Gives elastin its elasticity - makes it an interconnected rubber network

Marfan syndrome

- FBN1 codes for fibrillin

A person with which of the following oncogenes is most likely to develop cancer?

A. myc

B. ras

C. myc + ras

C. myc + ras

-there is cooperativity of oncogenes - a single mutation will not likely cause cancer

A protein tyrosine kinase

v-src (v=viral) is a constituitively active mutant of c-src

What do chromosomes look like during mitosis?

During G1?

during interphase?

mitosis - condensed "X"

G1 - threadlike

Interphase - occupy "territories" only visible by staining

- DNA is highly acidic (has a - charge) due to the phosphodiester bonds linking the nucleotides (PO4-)

- Histone proteins are basic (have a + charge) due to their basic residues of histidine, lysine, and arginine

What effect does methylation of DNA have on transcription?

Methylation of DNA represses transcription

What effects do acetylatioin and phosphorylation of histones have on transcription?

Both acetylation and phosphorylation of histones activate transcription.

 What are three ways that immune cell chromosomes differ from those of other cells?

(Characteristics that ultimately make them great for producing antibodies, but bad for cloning)

-Combinatorial joining of gene segments

-Imprecise joining of segments (junctional diversification)

-Somatic hypermutation

True or False:

Telomerase activity is often deactivated in cancer.

False

It is often activated in cancers.

Telomerase is required to keep telomeres large and robust.

What structure in the chromosome holds sister chromatids together and allows for attachment of the mitotic spindle during mitosis?

Centromere

Describe the anatomy of a nucleosome.

A nucleosome consists of double-stranded helical DNA wound around a core of 8 histone proteins (H2A, H2B, H3, H4 x 2), forming "beads on a string". The beads are linked together by H1 histone linker proteins.

What are HATs and HDACs?

HATs: Histone acetylases

HDACs: Histone deacytylases

-these are major chemotherapeutic targets

95% of the proteins found in the mitochondria are encoded by nDNA.

mtDNA codes for 13 proteins, which are synthesized in the mitochondrial matrix and function in electron transport.

1. maternal inheritance

2. heteroplasmy (wild type mixed with mutant)

3. stochastic (random) segregation of mtDNA

4. threshold effect (of mutations)

What 3 required proteins form the machinery for mtDNA replication ?

POLG - mtDNA polymerase g

mtSSB - mtDNA single stranded binding protein

Twinkle - mtDNA helicase

What cells are the primary target of HIV?

CD4+ T-lymphocytes

- CD4 receptors bind HIV gp120

HIV virus

(p24 is part of the virus capsule)

Where in the HIV RNA are the LTR regions located?

What do these regions do?

LTR (long terminal repeat) regions located at 5' and 3' ends.

-they are invariant, conserved domains

-serve as binding site for host transcription factors and allow for integration into host DNA

POLG

- coding for mitochondrial DNA polymerase

Mutations in these CD4 co-receptors would likely decrease their chances of infection and/or increase their chances of survival.

Which part of tRNA accepts amino acids?

Which part talks to mRNA?

The 3' CCA end is where amino acids are esterified and attached to the growing protein.

The Anticodon portion of Loop 2 talks to mRNA.

prokaryotes: FMet recognized by tRNA as START Met

eukaryotes: IMet recognized by tRNA as START Met

MetM is for internal Met in both euk & prok

- there is redundancy in the code such that 61 codons only codes for 20 amino acids

- also, there is WOBBLE pairing in the 3rd position which allows 1 tRNA to recognize more than one amino acid

Aminacyl-tRNA synthetase catalyzes the reaction of the amino acid with ATP, which activates it for transfer to tRNA.

These steps allow for proofreading and ultimately ensure the fidelity of protein synthesis.

What do Streptomycin, Tetracycline, Chloramphenicol, and Erythromycin all have in common?

How are they different?

-they are all antibiotics that target prokaryotic ribosomal subunits to inhibit protein synthesis

-Strep. and Tetra. target 30S, while Chlora. and Erythro. target 50S

-Strep. is Aminoglycoside,interferes w/ Initiation

-Tetracycline inhibits binding of AA-tRNA to A site

-Chlor. inhibits peptidyl transferase

-Erythro. is a Macrolide, inhibits translocation

Puromycin is a toxin that inhibits protein synthesis in both prokaryotes and eukaryotes.

It imitates AA-tRNA and causes premature release of nascent polypeptides.

Protein secretion is cotranslational!

secretion sequence (60-70 residues) is recognized by SRP (signal recognition peptide) in the cytosol during translation. SRP controls elongation and targets protein for maturation in ER.

When is trancription of lac operon mRNA high?

A. -lactose, -cAMP, +glucose

B. +lactose, -cAMP, +glucose

C. +lactose, +cAMP, -glucose

C. +lactose, +cAMP, -glucose

Lactose derepresses the constituitive repressor, allowing the polymerase to bind to the promotor region. cAMP needed for transcription.

RNA Polymerase III

in the nucleoplasm

CRP

(c-reactive protein)

-produce by liver in response to disease, trauma

-marker of systemic inflammation

Blue sclera is a sign of what disease?

OI

(osteogenesis imperfecta)

Which residue(s) are selectively glycosylated in procollagen?

Lysine , with glucose and galactose