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45 Cards in this Set
- Front
- Back
During replication, DNA polymerase initially incorporates 1 wrong nucleotide every ~106bp. However, the overall accuracy is much higher (1 error per 109bp). Why?
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Repair mechanisms are present to decrease the incidence of an error during replication
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In which direction is DNA being synthesized biosynthetically?
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5’ – 3’ direction. If replication occurred 3’ – 5’, could not proofread because no PPP.
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Double-stranded DNA is fairly stable and strand separation usually requires high energy. Instead of high temperature, how do organisms cope with this “problem” during replication?
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Through the use of enzymes
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What is an Okazaki fragment and in which process does it play a role?
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An Okazaki fragment is a short fragment of DNA created on the 3’ – 5’ lagging strand during DNA replication. It’s role is to allow the lagging strand to be replicated in the 5’ – 3’ direction.
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Name three activities of DNA polymerases.
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Polymerase (gap filling)
5’ – 3’ exonuclease activity (proofreading) 3’ – 5’ exonuclease activity ( primer removal, RNAse H) |
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Name a process in which DNA ligase plays a role.
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Ligation of Okazaki fragments
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Name a process in which DNA topoisomerase is important.
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Untwisting of parental strands in DNA replication
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Which functional group of DNA ligases and topoisomerases is thought to play a catalytic role?
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Phenolic hydroxyl group on the Tyr residue.
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What is the importance of the 5’ → 3’ exonuclease activity of DNA polymerase I in E. coli?
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Primer removal, RNAse H activity
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What is the difference between processive and distributive enzymes and name an example for each.
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Processive- carries out multiple nucleotide additions before disassociating Ex. Pol III
Distributive- readily dissociates from DNA Ex. Pol I |
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Does DNA synthesis occur in one or two directions from the ori?
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In both directions from the ori.
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Drugs that inhibit DNA synthesis usually cause cell cycle arrest at which phase?
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G1
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What are telomeres?
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TTAGGG repeats added to the ends of linear chromosomes to protect the ends from shortening and losing genetic information.
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Why is it advantageous that thymine rather than uracil is a DNA base?
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If U was used instead of T in DNA, it would increase the risk of mutation. C can easily be oxidized to form U. If this occured, then U would base pair with A instead of C with G which would result in unrecognized errors in transcription.
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Which promoter sequences are being recognized by bacterial RNA polymerase?
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-35 region and the Pribnow Box
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How can promoter sequences be identified?
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The -35 region and Pribnow box are both highly conserved regions that the promoters can recognize.
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Where does the σ factor play a role?
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Specific σ factors recognize classes of genes in Prokaryotes. Along with a Polymerase they bind to the promoter to initiate transcription.
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Name a characteristic of high promoter efficiency.
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Several RNS polymerases can initiate transcription simultaneously.
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How is it ensured that transcription occurs in one direction?
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The -35 region is asymmetrical and the enzyme can recognize the direction of transcription.
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What is a difference between DNA and RNA polymerase with respect to the necessity for primers?
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DNA requires a primer, RNA does not require a primer.
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Which nucleoside analog do you expect would inhibit elongation: 3’-deoxyadenosine or 5’-deoxyadenosine and why?
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3’- deoxyadenosine because it is missing the PPP (triphosphate)
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Which process do intercalating chemotherapeutics interfere with?
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They interfere with replication and transcription.
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What is the basis for ρ-independent termination of transcription in prokaryotes?
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GC rich palindrome upstream of 6-7 U (A on template DNA). RNA forms a stem-loop secondary structure which results in termination.
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Would the use of ITP instead of GTP during in vitro transcription increase, decrease or not affect termination efficiency?
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ITP would diminish termination efficiency because I:C pairs are weaker than G:C pairs (2 vs. 3 H bonds)
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What is the TATA box?
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Promoter sequence for transcription in Eukaryotes.
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What are common characteristics of transcription factors?
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Contain both DNA recognition helix and protein interaction domains
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What is a Holliday junction and in which process is it thought to be an intermediate?
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o Homologous recombination
o Holliday junction formed from 4 DNA strands o Forms Holliday intermediate |
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What is a heteroduplex in homologous recombination?
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o Regions where one strand was from one duplex, the other from the homologous duplex; caused by reciprocal inversion
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How can you recognize a transposon in the genome?
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o Insertion sequence has inverted repeats at both ends
o Insertion sequences are flanked by direct repeats in the same orientation in target sequence |
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Name different types of DNA repair processes.
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o Direct repair
Cyclobutane thymine dimmer photoreactivation • Formed by [2+2] cycloaddition • Photolyase binds • Light absorption (300-500nm) cleavage • Photolyase dissociates • Demethylation Demethylation- alkyltransferases (methyl guanine methyl transferase) reverse this process o Excision Repair Base Excision Repair • Part of DNA excised by DNA glycosylase • Removal of damaged base by specific glycosidases • Repair of AP sites left after removal of damaged base (or by spontaneous hydrolysis); o AP endonucleases & lyases excise abasic sugar o Gap filling (DNA polymerase, pol o Ligation (DNA ligase) Nucleotide excision repair • Excises several nucleotides • Excision repair of photodimers • Endonucleases excise • Gap filling by DNA polymerase • Ligation |
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o Special Cases of Nucleotide Excision Repair
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Transcription coupled repair- only occurs if you need proteins around
• Lesions also block transcription • Transcribed regions are repaired faster Mismatch repair • If wrong base is incorporated (undamaged) |
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What is the importance of DNA methylation in DNA repair?
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o How to recognize newly synthesized strand (and not “repair” parental strand)?
A: Look at methylation pattern! Higher degree of methylation in parental strand. |
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Sometimes damaged DNA is not repaired yet by direct or excision repair when the replication complex approaches a lesion. Which repair mechanism(s) can come into play?
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o Translesion synthesis, recombination repair by daughter strand gap repair, double-strand break reapir, or repair of replication forks
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What are bypass polymerases, what statement can you make about their processivity?
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o Bypass polymerases are only induced upon extensive DNA damage (SOS response) and readily dissociates from DNA (distributive)
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Does coupled transcription-translation occur in prokaryotes eukaryotes, or both?
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o prokaryotes
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What statement can you make regarding the degree of RNA modification in prokaryotes versus eukaryotes?
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o More dramatic
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Name features of eukaryotic mRNA.
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o 5’-terminal cap
o 3’-terminal poly(A) tail o Methylated internal variants o Splice variants |
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Name 3 unique structural features of tRNA.
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o Nucleosides are highly modified
o Anticodon o 3’-CCA acceptor arm for aminoacylation o many non-Watson-Crick base pairs |
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What is splicing?
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o excision of intron sequences
o Creation of several splice variants possible ie. Exon between two introns-> modified functionality ie. Depending on cell type o Increased protein diversity relative to prediction from genome, more variability for organism o Other ways to generate protein diversity: posttranslational modifications |
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Why could not DNA be spliced by the mechanism discussed in class?
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o No 2’-OH
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What is a putative advantage of splicing?
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o Creation of several splice variants possible- modified functionality depending on cell
o Increased protein diversity relative to prediction from genome |
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Which sequence is for the attachment of amino acids onto tRNAs?
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o CCA required for attachment of amino acids
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Why is rRNA originally transcribed as one unit?
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o want unit 1:1:1 ratio (of the S segments)
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How can you distinguish prokaryotic from eukaryotic rRNA?
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o Prokaryotes
28S rRNA 5.8S rRNA 18S rRNA o Eukaryotes 23S rRNA 16S rRNA 5S rRNA |
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What are small interfering RNAs?
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o 21-23 ribonucleotides long
o Facilitate sequence specific gene suppression via RNA interference (of homologous genes) |