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247 Cards in this Set
- Front
- Back
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What is the criteria for the time frame in treating HIV?
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The earlier the better, because at a certain point the virus replicates heavily and destroys all CD4+ cells, which is difficult to reverse
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What is an ideal target for antiviral therapy?
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Viral enzymes or proteins that don't have a cellular counterpart
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What do development of assays help with?
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Allows/facilitates the screening of large libraries of compounds that can inhibit the enzyme.
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What is Phase 1 of clinical trials?
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Test drugs at different doses on healthy volunteers, for toxicity effects
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What is Phase 2 of clinical trials?
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Test drug potency (efficacy), and the dose that has the best efficacy
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What is phase 3 of cliinical trials?
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A large trial to see for complications and long-term effects
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Which generation of drugs are the most vulnerable to development of resistance?
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The first generation
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What are set points?
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Looking at the level of CD4+ cells and the viral load, to decide when to begin treatment
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What is AZT?
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A monotherapy, a primary nucleoside inhibitor, but resistance develops quickly
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What are drug cocktails generally composed of?
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Protease inhibitors, reverse-transcriptase inhibitors
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What is the crystal structure like of reverse transcriptase inhibitors?
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A human right hand
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What are two types of RT inhibitors?
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Nucleoside analogue inhibitors, or non-nucleoside analogue inhibitors
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What are nucleoside analogue RT inhibitors effective against?
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HIV-1 and HIV-2
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What are some examples of nucleoside analogue RT inhibitors?
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AZT, d4T, 3TC
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What is a common feature of all nucleoside analogue RT inhibitors?
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They all lack the 3' OH, which what is normally attacked by an incoming nucleotide. Therefore they act as CHAIN TERMINATORS!
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What are non-nucleoside analogue RT inhibitors active against?
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HIV-1, but NOT HIV-2 (unlike NRTI's)
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How do NNRTIs work?
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They bind to hydrophobic pockets away from the active site
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What is Atriola?
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A once a day drug that combines nucleoside and non-nucleoside inhibitors
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What is an example of a protease inhibitor?
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Indinavir
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How do protease inhibitors act?
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Bind to the active site and prevent the substrate from binding and being cleaved by the protease
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How does Ritonavir (a protease inhibitor) act?
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Mimics the precursor peptide being cleaved
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What is a "sustained biological response"?
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When the viral load decreases quickly and stays down
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How does administration of Ritonavir affect other protease inhibitors?
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It enhances the potency of other protease inhibitors when co-adminstered with them
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What is the difference between anti-HCV and anti-HIV therapy?
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Eradication of HIV is not possible as it is integrated into the genome, while eradication of HCV is.
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What is the standard of care for HCV?
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Pegylated interferon-alpha and ribavirin
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What is a negative aspect of drug-drug interactions? (drug cocktails)
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For instance, when using two nucleoside analogue inhibitors with ribavirin, they can reduce each others levels in the cell!!
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What is non-adherence?
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If a patient doesnt respond to a drug right away then the drug levels will fluctuate and they wont be able to block replication, leading to mutated viruses and resistance
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What is HAART?
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Highly active antiviral therapy- used to reduce plasma RNA levels of HIV below the level of detection
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What is pharmacokinetics?
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What the body does to the drug
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What is the difference between Genotypic and Phenotypic resistance assays?
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Phenotypic just tells us if a virus is resistant to a drug or not, genotype reveals mutations (which might proceede the phenotypic resistance)
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What are other drugs that can be used to target HIV?
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Fusion inhibitors,
Entry inhibitors HIV integrase strand transfer inhibitors |
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What is the key difference between Hepatitis B and retroviruses?
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Retroviruses contain an RNA genome and HBV conains a partially double stranded DNA genome
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What are the general properties of retrovirus genomes?
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Postive strand RNA, 7-10kb, when packaged exists as a dimer
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Which gene products can be found inside the viral capsid?
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The pol gene products (integrase, reverse transcriptase, viral protease)
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What is the surface protein of retroviruses?
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gp120
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What is special about the HIV glycoprotein gp160?
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It is cleaved by cellular proteases (NOT VIRAL)
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What is the transmembrane protein of HIV?
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gp41
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What is the viral infectivity factor (Vif) of HIV for?
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Fights cellular factor APOBEC
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What does the cellular factor APOBEC do?
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Restricts the infection of the HIV virus
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What are two viral proteins that HIV can live without?
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Nef and Vpr
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What is Nef for?
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When replication is diminished, Nef is the major contributer to development of pathogenesis
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What is the first interaction upon HIV entry?
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Surface glycoprotein gp120 with CD4+ receptors on the CD4 + T lymphocyte
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What does the interaction of gp120 and CD4+ receptors result in?
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The exposure of the transmembrane protein gp41
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What does gp41 interact with?
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Co-receptors, chemokine receptors CC5R5 or CCR4
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What happens after the interaction of gp41 with the co-receptors?
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Ultimately the membranes are fused and a pore is generated that the viral capsid can pass through into the cell
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Once the dsDNA generated from reverse transcription goes into the nucleus, where does it tend to go?
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To areas that are actively transcribed, where their chromatin are not so condensed and are more open
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What performs transcription of the integrated dsDNA?
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Cellular RNA polymerase II
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How well does cellular RNA polymerase II perform the transcription to viral RNA?
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It is not very efficient and tends to abort early.
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How often can the cellular RNA pol II generate full length mRNA in transcription of integrated dsDNA?
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Only 1 out of every 100 transcription events
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How does HIV ensure that it will always produce the full length mRNA?
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Its mRNA generates the TAT protein
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What does TAT do?
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Goes back into the nucleus and binds to the 5' Tar region of the viral RNA
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Where is the 5' Tar region located?
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In the 5' non-translated region
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What does the TAT protein do once it is bound to the Tar region?
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Recruits cellular factors, including a cellular kinase
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What does TAT recruitment of the cellular kinase allow?
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Phosphorylation of the RNA polymerase, making it more processive so that full length RNAs are generated all the time
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How does splicing work?
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The 2' OH of the intron attacks the 5' phosphate, resulting in exon 1 leaving. The 3'OH of exon 1 now attacks the junction between intron and exon2, and this results in the joining of the two exons and the intron leaving as a LARIAT
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What happens once lots of full length viral RNAs are generated?
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The virus wants to stop splicing so that it can generate longer RNAs that encode for structural proteins, so that they can encapsidate all these viral RNAs to make progeny virions
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What is the purpose of the Rev protein?
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Rev goes back to the nucleus once it is generated in the cytoplasm, and binds to Rev Responsive Elements (RREs)
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What does Rev binding to RREs do?
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Diminishes splicing and favors the generation of unspliced (encodes for Gag and Pol) and singly spliced (encodes for Env) mRNA, resulting in the ability to generate the STRUCTURAL proteins!
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What is the use of Vpr?
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Is involved in the pre-integration complex
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What are the early stages in the HIV life cycle?
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Attachment to integration
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What are the late stages in the HIV life cycle?
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Transcription to viral particle release
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What type of splicing results in the generation of the early phase proteins?
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Highly spliced RNA
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What type of splicing results in the generation of the late phase proteins?
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Not as much splicing
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What are the "early proteins" generated?
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Tat, Rev
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What are the "late proteins"?
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Env, Vpr, Gag, Pol
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How are the Gag proteins arranged?
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In one ORF that generates a Gag polyprotein, that is cleaved by VIRAL proteases at later stages
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What is the Gag ORF composed of?
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4 proteins with spacer peptides
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What are the four proteins in Gag?
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-MA (Matrix/membrane associated) (p17)
-CA (capsid, p24) -NC (nucleocapsid, p7) -p6 |
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What are gag proteins involved in?
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The assembly process
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Where is the matrix protein found?
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On the inner side of the membrane, forms a membrane bound layer
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What component of MA is required for membrane binding?
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The N-terminal myristyl group is required for membrane binding
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What does the myristyl group of MA attachment to the membrane allow?
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Directing of all the other viral components to the inner side of the membrane, where assembly process can take place
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How is gag targeted to the membrane?
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By the matrix domain!!! (because it is myristylated!!!)
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What is the role of the capsid protein?
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It has a role in particle assembly
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What type of structure does the capsid form?
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A conical core, with hexamers. THere is a hole in the centre that allows nucleotides to enter
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What happens to the capsid when HIV infects a new cell?
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The capsid structure is released into the cell, the conical structure remaining intact. It only is destroyed after the reverse transcription process
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What is the purpose of the nucleocapsid protein?
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Serves as a nucleic acid chaperone
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What are some features of the nucleocapsid protein?
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It has a highly basic N terminus and zinc fingers that bind HIV RNA
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How does NC exert its chaperone effects?
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-NC unwinds the tRNA primer, as well as helps place it on the PBS. Consequently, NC facilitates processive DNA synthesis!
-NC stabilites the RT enzyme -NC interacts with Vpr |
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How are Gag and Pol organized?
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In different reading frames
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How is Pol generated?
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By first creating the Gag-Pol precursor!
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What is the only way the Gag-pol precursor can be generated for HIV?
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Through a ribosomal frameshift (-1)
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What are the pol proteins?
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Protease, reverse transcriptase, RNaseH, integrase
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What type of protein is RT?
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A heterodimer, with p51 subunit and the Nterminus of the p66 subunit
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What does the C terminus of the p66 subunit of Pol represent?
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RNasH
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Which processes in the HIV life cyce require cellular factors?
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Transport of the dsDNA into the nucleus and integration into the host chromosome
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What are the two "reactions" of integration and where do they occur?
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1) 3' processing (in the cytoplasm)
2) Strand transfer (in the nucleus) |
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What occurs in 3' processing?
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The viral integrase chops off the 2 terminal GT nucleotides from either end of the viral DNA
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What is this 3' processing reaction catalyzed by?
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A water molecule that acts as a nucleophile
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What does the 3' processing result in?
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Exposure of new OH groups at the 3' viral ends
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How does the generation of the new 3'OH groups help with the strand transfer that occurs in the nucleus?
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The 3'OH groups act as nucleophiles and can disrupt internucleotide phosphodiester bonds of the target DNA, resulting in the covalent linkage of the viral DNA within the host genome
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Where do cellular factors come in to play in the integration reactions?
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Once the strand transfer is complete, there are still gaps which are fixed by the CELLULAR REPAIR MACHINERY
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What do current integrase inhibitors target?
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The strand transfer event (mediated by viral integrase)
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How does integrase bind the ends of DNA?
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As a dimer!!
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What type of proteins does IN recruit?
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Vpr, NC, cellular factors
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Which cellular factor helps bring proviral DNA to the chromosome?
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LEDGF/p75
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What is LEDGF?
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Lense epithelium-derived growth factor/transcriptional co-activator 75
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What family is LEDGF apart of?
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HDGF (Hepatoma-derived growth factor)
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Which portion of IN interacts with LEDGF?
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The portion that contains the catalytic domain
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Where does LEDGF interact?
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At the interface of the two integrase monomers is a groove where a portion of LEDGF can bind! (known as IBD (integrase binding domain))
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How does LEDGF function?
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It mediates chromatin binding of HIV pre-integration complexes. It does so by its N terminal sequence elements, by tethering HIV-1 PIC to chromatin for integration
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What is Raltegravir?
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Integrase inhibitor
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What are "host restriction factors"?
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Host factors that FIGHT viruses
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What is an example of an innate immune response to block viral replication?
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Interferon response
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What are two key cellular factors that act as restriction factors against HIV?
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TRIm-5alpha, and Human APOBEC3G
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Which family is TRIM-5alpha a member of?
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Tripartite Motif Protein Family
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What does Trim-5alpha prevent?
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Lung cells from being infected by HIV
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How is the TRIM-5alpha genome organized?
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In 3 domains
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What is one of the key domains of TRIM-5alpha?
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A ubiquitin-ligase substrate specificity
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How does TRIM-5alpha use this ubiquitin-ligase substrate specificity domain to combat HIV?
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It is thought that since TRIM-5alpha can bind the capsid proteins of HIV< that it can recruit the ubiquitin machinery and direct the viral particle to the proteosome
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What are some other theories as to why TRIM-5alpha restricts HIV?
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-TRIM5a complex can trigger innate immune responses
-TRIM5a can coat the viral particle and disrupt core rearrangement/uncoating |
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Is human TRIM-5alpha effective in restricting HIV?
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No, only Rhesus macaque orthologs have been seen to be able to restrict HIV in human cell lines
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How was APOBEC3G discovered?
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By working on viral protein Vif
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What is viral protein Vif necessary for?
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In the virus-producing cell processes following post-entry (ie. steps before reverse transcription, and PIC entry)
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How do Vif and APOBECE3G interact?
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Vif counteracts the antiviral activity of APOBEC3G
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How does APOBEC3G act?
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By editing the RNA of the virus, and as a result diminishing its replication
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How does APOBEC3G edit the RNA?
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APOBEC posseses a DEAMINASE activity, converting a cytosine to a uracil, resulting in the incorporation of an A in the 2nd DNA strand, instead of a G. Therefore it generates a
G-> A mutation. |
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Where is APOBEC3G found?
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Incorporated into viral particles, or in the cytoplasm
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Why does the G-> A mutation by APOBEC3G result in a negative effect on HIV?
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RT will incorporate the wrong nucleotide in a critical area and the enzyme will drive the virus to catastrophe (death)
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What is a late stage HIV restriction factor?
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Tethrin/CD317
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What type of protein is Tethrin/CD317?
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A membrane bound protein
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What HIV protein counteracts Tethrin/CD317?
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Vpu, by interacting with tethrin in a way leading to its internalization and proteosomal degradation
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Describe the progression of a typical HIV infection
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-The primary infection shows a dramatic decrease in CD4 cells, as well as a dramatic increase in viral load (viral RNA)
-A latency period is established -Onset of AIDS symptoms, correlating with a sharp drop in CD4 levels |
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What does AZT have instead of the 3'OH group?
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C-N3
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What does d4T have instead of the 3'OH group?
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C-C double bond
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What does 3TC have instead of the 3'OH group?
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An S replacing the C that the 3'OH is normally bound to!
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What are AZT and d4T analogues of?
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Thymidine
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What is 3TC an analogue of?
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Cystidine
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What are Efavirenz and Nevirapine?
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Non-analogue RT inhibitors
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How do Efavirenz and Neviparine act?
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Bind to hydrophobic site, causing a reduction in the effectiveness of the active site
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What is a requirement for the functioning of analogue inhibitors?
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The nucleosides need to be phosphorylated (sometimes by cellular kinases)
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Why is acyclovir normally inefficient against HIV?
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Because acyclovir requires activation by phosphorylation
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How is acylcovir able to be used against HIV?
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If the patient with HIV is co-infected with Herpes Simplex Virus, the HSV provides the kinase that is required to activate acyclovir
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What is combivir?
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A combination of 2TC and AZT
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What is Atripla?
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A single pill that has three drugs
(non-nucleoside analogue inhibitor (Efiavirenz) and two nucleoside analogue inhibitors (Tenofovir, emtricitabine) |
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What is Ritonavir?
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A Protease Inhibitor Boosting drug, binds to enzymes and cytochromes that would otherwise degrade the protease inhibitor, preventing its function
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When considering virus population, when should a drug be administered and why?
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When the viral load is still small, as overtime the virus will generate mutations and be resistant to the drug!
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How are mutations in HIV generated?
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RT has low fidelity, creates 1 mistake per genome, and has no proofreading
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What are the last stages of HIV related disease characterized by?
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High levels of replication, mutations and exponential viral growth
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What is an example of a mutation that causes a resistance for 3TC?
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The methionine at pos 184 (M184) can undergo two mutations M184I or M184V
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What is the M184I mutation?
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ATG to ATA, resulting in a isoleucine codon
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What is the M184V mutation?
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ATG changes to GTG, resulting in a valine codon
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Which mutation has a higher replication capacity and results in overpowering the other in the population of HIV?
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M184V
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What does the K103N mutation confer resistance to?
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All non-analogue RT inhibitors
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What does Q151M mutation confer resistance to?
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All nucleoside RT inhibitors, except 3TC
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While M184V confers resistance to 3TC, what other effect does it have?
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It increases the susceptibility to AZT! (Another nucleoside RT inhibitor!)
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What does this resistance induced by M184V allow?
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People can use two drugs to select for different, incompatible resistance pathways
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What is the AZT resistance mechanism?
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RT incorporates AZTmonophophate, but AZT can be excised by pyrophosphate
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Where does the pyrophosphate come from?
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ATP acts as the donor of pyrophosphate
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What is better genotypic or phenotypic determination of resistance?
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Genotypic, which involves sequencing and comparing mutations to known mechanisms of resistance is less costly and time consuming than phenotypically testing viruses with different drugs and observing resistance
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What is Entecavir?
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Guanosine analogue that contains a 3' OH group, used to nihibit HBV
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How can Entecavir be used to treat HIV?
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If the individual is coinfected with HIV and HBV, HIV RT will incorporate ETV monophosphate
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What are three possible mechanisms of ETV drug action?
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De factor chain termination, delayed chain termination, or base pair confounder
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Which mutation induces resistance/lower efficacy of Entecavir?
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M184V
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Which mechanism is thought that ETV works by?
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Delayed chain-termination mechanism
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What are the two mechanisms of resistance for nucleoside analog inhibitors?
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Discrimination and excision
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What is "discrimination" in this sense?
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The mutations in the reverse transcriptase enzyme discriminate against the inhibitor in favor of the natural nucleotides
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Where are all the mutations that confer resistance based on discrimination located?
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These mutations are all clustered around the nucleotide binding/active site
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What are some mutations that lead to discrimination?
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K65R, M184V, L74V, Y115F
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What is K65R close to?
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The phosphate moiety
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What does K65R confer resistance to?
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Tenofovir, as well as 3TC
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Where is the M184V mutation located?
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Near the sugar
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How do drugs perform their resistance mechanisms on AZT?
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It is excised
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Why are mutations resulting in resistance to AZT not directly clustered around the active/nucleotide binding site?
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Because these mutations facilitate the binding of ATP in such a way that it can attack AZT (that is in the nt binding site)
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What are TAMS?
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Thymidine analog-associated mutations (mutations that are slightly distant from the active site)
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What are two drugs that select for TAMS?
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AZT and D4T
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Which drug is not very well excised?
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3TC
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How do M184V viruses increase susceptibility of the virus to AZT?
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TAMs increase the excision of incorporated AZT monophosphate, but M184 diminishes this reaction
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Why isnt it a good idea to use three nucleoside analogue RT inhibitors instead of two in a drug cocktail?
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1) Pharmacokinetics: these drugs in a cocktail may not get sufficiently phosphorylated
2) K65R- if we combine certain drugs, this mutation will show up and many drugs will encounter resistance |
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Which mutation confers resistance to all NRTIs?
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Q151M
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What type of mutation is that of Q151M?
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It is not sufficiently fit and the replication capacity of the virus will go down.
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Which generation of the virus can contain mutations that counteract the efffects of the resistance conferring mutations?
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3rd or 4th generation
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Where can the 3rd generation mutations be located?
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Outside the RT enzyme or just different regions from the nucleotide binding site
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What are some examples of older drugs that have mutations in the area not corresponding to the primer template?
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AZT, Niverapin
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What allows RT to sit on the template for longer periods of time, ensuring removal of nucleotide?
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The mutations that are located distant from the active site also diminish the RNaseH activity, allowing the template to be around longer
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What is a fusion inhibitor drug?
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Fuzeon (enfuvirtide, T-20)
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What is an entry inhibitor?
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Selzentry (maraviroc)
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What is an HIV integrase strand transfer inhibitor?
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Isentress (Raltegravir)
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What is the problem with using maraviroc?
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It is an antagonist for the CCR5 co-receptor. However, this just selects for virus strains that use CXCR4 as a co-receptor
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How did they prove that Entecavir (an HBV drug) can be effective against HIV?
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They saw that the M184V mutation in the RT conferred resistance to it
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What type of drug is Entecavir?
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A guanosine analog that contains the 3'OH group
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Since the 3'OH group is present, how does Entecavir act as a chain terminator?
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yes, The hydroxyl group is not properly aligned to attack the alpha phosphate.
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What are other mechanisms of Encetavir action?
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-Delayed chain terminator (a few nt can be added and then stops)
-Base-pair confounder (Entecavir enters the nascent DNA strand, and the daughter strand is complete. But when the DNA strand containing ETV is used as a template, problems arise) |
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Which mechanism of Encetavir action is dominant and what does this help with?
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Delayed chain terminator, helps the analog from being excisied
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What is the proposed mechanism of the prevention of ETV excision?
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When ETV is at n+3, it prevents RNaseH activity of RT from making one of its three cuts by repeling the active site of the polymerase.
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Where does Hepatitis C replicate?
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On the inner side of the ER
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What is STAT-C?
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Specifically targeted antiviral therapy for HCV
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What are the two most promising targets in treating HCV?
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The protease (NS3) and polymerase (NS5B)
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What is the difference between the HIV protease and the HCV protease?
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HCV protease (NS3/4A) acts on a single polyprotein that needs to be cleaved
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What is telaprevir?
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An HCV protease inhibitor
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How can protease inhibitors indirectly increase innate immunity against HCV?
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Since viral protease can interfere with signal transduction of an interferon response, once this protease is inhibited, this interference no longer occurs!
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What is the problem with using protease inhibitors along with ribavarin and interferon for some patients?
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Some patients simply do not respond to ribavarin and interferon, so they are basically on a "monotherapy" of protease inhibitor, and resistance can develop quickly
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What is boceprevir?
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An HCV protease inhibitor
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Why do we use non-nucleoside analog inhibitors to treat initiation of RNA synthesis by HCV NS5B, and nucleoside analog inhibitors to treat elongation of RNA synthesis?
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Polymerase initiation is fragile, while its elongation is stable/robust
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Why cant we use obligate chain terminators in HCV?
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Because NS5B incorporates ribonucleotides, which are non-phosphorylated and lack the 3'OH group
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Do inhibitors of HCV NS5B act as delayed chain terminators?
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No because other modifications in these inhibitors prevent the next nucleotide from binding to the complex. Therefore, inhibitors of HCV NS5B act as non-obligate chain terminators
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What are the four sites that HCV NS5B inhibitors can bind?
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NNI site A/ Thumb1
NNI site B/Thumb2 NNI site D/ Parm 2 NNI site C/ palm 1 |
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Which drugs bind to NNI site A?
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Benzimidazoles
Indoles |
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Which drugs bind to NNI Site B?
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Pyranoindoles
Thiophene Dihydroxpirones |
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Which drugs bind to NNI site D?
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Benzofurans
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Which drugs bind to NNI site C?
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Thiadiazines
Acyl-pyrrolidines |
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What is a key difference in the HCV polymerase structure, and the HIV RT structure?
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HIV RT is open, while HCV pol is closed
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How do inhibitors of the HCV Ns5B prevent it from forming its closed structure?
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If the inhibitor is present on the polymerase, it prevents the conformational change where the finger domain can fold over and interact with the thumb domain to trap the RNA
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What are some differences of HIV with HCV pol?
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HIV-RT NNRTIS can bind and work at every step
There is only 1 binding site on HIV RT |
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What is the only class of compounds that can inhibit all genotypes of HBV?
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Nucleoside analog inhibitors, however these are more TOXIC
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What family does HBV belong to?
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Hepadnaviridae
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How is HBV transmitted?
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Through blood and sexual contact
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What type of disease does HBV cause?
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Chronic, mostly liver disease
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What is the genome like of HBV?
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3.2kbp, circular, partially dsDNA
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What is a unique feature of HBV?
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There is a reverse transcription step of pregenomic RNA
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What shape does the HBV core protein have?
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Icosehedral
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How many envelope proteins does HBV have?
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Three
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How are HBV vaccines designed?
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Using anti-HBs (the surface antigen of the HBV viral envelope). They are designed using the PreS/S region of the small surface protein
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What are some ways to treat HBV?
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Interferon, 3TC or Tenofavir
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How does HBV entry occur?
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Receptor mediated, injects genome/core into cytoplasm
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What happens once the virus enters the cell?
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The partially dsDNA enters the nucleus to be transcribed to RNA
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What are two major differences of HBV and HIV?
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There is no integration of the HBV genome in the host, and the reverse transcription process of HBV occurs POST ENCAPSIDATION!
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What does the virus use cellular machinery for?
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To produce RNA, transport it to the cytoplasm, and initiate translation
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How is viral RNA attached to the core for encapsidation?
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Viral RNA forms a hairpin epsilon structure, which allows the attachment
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Where does maturation of HBV viral particles occur?
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At the ER/Golgi
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How do HBV particles exit the host cell?
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By exocytosis
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Where does HBV viral particle uncoating occur?
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In the cytoplasm
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What happens once HBV is uncoated?
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The nucleocapsid is transported to the nuclear membrane and the partially dsDNA circular genome is injected into the nucleus
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What happens to the partially dsDNA genome once the virus enters the cell?
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1) The plus strand is extended by viral DNA pol, using RNA primers
2) The P and RNA primer are removed by host cells enzymes 3) The resulting 3'OH and 5'PO4 ends are ligated by cellular enzymes |
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What is special about the HBV polymerase?
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It does not only contain RT activity w/ RNAseH domain, it also has a terminal linker domain that is required for RT
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What happens once the full circular dsDNA genome is generated?
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There are 2 regulatory elements, direct repeats DR1 and DR2, which play a role in reverse transcription
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What are the types of mRNA generated from the HBV genome?
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-Long, pre-genome RNA (pgRNA)
-PreS1 -PreS2 -X mRNA |
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What does pgRNA encode for?
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The core protein
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What does preS1 encode for?
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Large surface protein on the envelope
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What does preS2 encode for?
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Middle and small surface proteins on the envelope
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What does the X mRNA encode for?
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"Mysterious" protein X, implicated in transcription, and in inducing cancer and other functions
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What are the four different ORFS of the HBV genome?
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1) Core C and Pre-core (2mRNAS)
2) PreS1, PreS2, S (3mRNAs) 3)Polymerase (P) -encoded by pgRNA 4) X protein |
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What are Enh1 and Enh2?
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They are enhancer elements that regulate transcription levels by allowing cellular transcription factors to bind
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What type of transcription factors bind Enh1 and Enh2?
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Hepatic cell specific factors, like Hnf1, Hnf3 and Hnf4
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What type of polymerase is encoded by the virus?
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A DNA polymerase that is RNA and DNA dependent
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What is a key difference in HIV and HBV initiation of DNA synthesis (reverse transcription)?
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Reverse transcription in HIV is primed by tRNA, and in HBV it is primed by a tyrosine (96) in a long region of the HBV gene products.
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How does the P protein bind?
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With its tyrosine to the epsilon sequence of the 5' end of the genomic RNA, by nucleophilically attacking the hydroxyl
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What occurs at the beginning of DNA synthesis?
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Four nucleotides are added and a STRAND TRANSFER event occurs
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Where does the strand transfer occur?
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The P + four nucleotides transfer to a homologous region at the 3' end
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What is the difference between the first strand transfer in HIV and HBV?
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In HIV, the template RNA is degraded by RNAseH, and this INITIATES the transfer. while in HBV there is no degradation by RNaseH until AFTER the transfer
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What happens after the first strand transfer?
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Most of the RNA template is degraded except for a small region which serves as an RNA primer
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What is the different between the degradation of the RNA template in HIV and HBV?
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In HIV, the ppt is resistant to degradation, but this is not the case in HBV
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What are the two possibilities to create the second strand of DNA?
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Either the second strand is generated straight from the RNA primer, or the primer undergoes a strand transfer to a zone of homology from DR1 to DR2 (at the 3' end)
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Which possibility creates the partially dsDNA and how?
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After strand transfer of the RNA primer, after about 50-80% of the 2nd strand has been synthesized, it just stops
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What is the difference in polymerase packaging in HIV and HBV?
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In HIV, 50-100 RT molecules are packaged, and in HBV only 1 is (but since it is covalently attached this is fine!!)
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What confers the genetic variability in HIV?
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RNA pol II transcription mechanism in HIV
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What confers the genetic variability in HBV?
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RT process
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