DNA replication Flashcards
Watson and Crick proposed structure of DNA allowed the molecule to be ____. Each parent strand serves as the ____ to line up the bases for the synthesis of a new daughter strand. Each new DNA helix consists of one strand from the parent helix and one newly synthesized ____strand
replicated. Template. daughter
The structure of DNA accounts for ___ and ___ of genetic information
storage; transmission
Meselson and Stahl proved that DNA replication is semiconservative:
One strand stays the same and serves as a template for the other strand. Other strand is made via complementary pairing of incoming nucleotides to the parent template strand.
How did Meselson & Stahl prove that DNA replication is semiconservative?
Grew E. coli for many generations in media containing N15 heavy isotope. They then transferred cells to N14 media and grew for successive generations and took sample for each generation time (cell doubling) and analyzed DNA density.
What was Meselson and Stahls technique?
Before transfer to media DNA had a high density. DNA after 1 gen had a density exactly in bet all N15 and all N14. After a second cell div. half of the DNA is all N14 labeled and half has the intermediate density which rules out non conservative replication which would have given all the DNA at a density 1/4 bet light and heavy
What is Kornberg known for
First to isolate DNA polymerase I from E. coli from nucleotide triphosphate precursors: dGTP, dATP, dCTP, and dTTP
what is Cairns known for
found out that replicating DNA has an extra loop (theta). DNA replication is bidirectional!
DNA polymerase exhibits processivity. What is processivity?
how long it stays on the template and adds bases before DNA polymerase enzyme dissociates from the template.
Polymerization is ____favored: hydrolysis of PPi and base pairing + stacking
thermodynamically
DNA polymerases are very ____. Sev mechanisms reduce error rate, such as mismatch repair mechanisms
accurate
how to errors in replication occur
protons form abnormal arrangements of e- that change the pairing ch’s of specific bases.
What is the mechanism of recognizing a tautomer
proofreading nuclease moves backwards and cuts base out. Polymerase resumes
DNA polymerase I has a ___function. DNA polymerase II has a specialized repair function. DNA polymerase III is the main DNA _____enzyme.
repair; replication.
DNA pol III has ___ subunits and ____ processivity
> 10; 500,000
What are the 3 stages of DNA replication
- Initiation (most complicated)
- Elongation
- Termination
DNA replication begins at the _____. It has DUE that has ____bp seq element which unwinds and form ss pieces that can be stabilized and start process of replication.
ORI; 13
THere are ____ R regions which are binding sites for DNA a protein which constrains the DUE. This causes it to open up into ss portions.
5
what are the proteins required to initiate replication at the E. coli origin
- DNA a: recognizes ori
- DNA b: helicase that unwinds DNA
- DNA c: Req for DNA b binding at origin (helper protein)
- HU: histonelike protein
- DNA G: synthesizes RNA primers. (Primase)
- SSB: binds single stranded DNA
- DNA gyrase: relieves torsional strain gen by DNA unwinding
- Dam methylase: methylates 5’ GATC seq at ori
what is the lagging strand
makes DNA seq piece meal in Okazaki fragment
How does replication initiate
- dnaA binds to oriC and forms initial complex
- dnaB/dnaC dimers bind to open complex and unwind origin DNA to expose ss template.
- ss DNA stabilized by binding of SSB’s
what is considered the prepriming complex
dnaA dnaB/dnaC
all synthesizing Dna polymerases require a ____ and can only add to the ____ end
primer; 3’ end of an existing strand.
____binds to dnaB/dnaC on DNA template. Other proteins add to form complex called the _____. Then a ______ dimer now binds to primosome to form the replisome. Leading strand synthesis begins at each fork using one DNA pol III molecule, the RNA primer and the leading strand template.
Primase; primosome; DNA pol III
what is primase
a specialized RNA pol that synthesizes a short 15 nt strech of RNA on DNA template which remains base paired.
DNA polymerases add bases to ____ hydroxyl end
3’
Physical dir of movement of replication fork on one strand in same direction as synthesis which is:
5’ to 3’
The new daughter strand is the ____strand. physical direction in the opposite direction to synthesis is for the other daughter strand.
daughter
solution to leaving long stretches of ss DNA is to syntheize ____strand discontinuously in short stretches called okazaki fragments
lagging
____ ____ and topoisomerase I maintain the chromosome negatively supercoiled.
DNA gyrase
how is elongation initiated
lagging strand is looped around on replisome so small portion of it is in same orientation as the leading strand which is how we get primers added on.
DNA polymerase I and ____ ___ repair the okazaki fragments and produce a discontinuous DNA lagging strand
DNA ligase
how does dna replication terminate?
Replication forks meet. Topoisomerases allow two double helices to sep
How is DNA replication in eukaryotes?
much more complex. Have 5 polymerases instead of 3
Whats the diff between bacterial and human DNA replication
- Linear chromosomes
- Multiple ori’s
- more DNA polymerases
what are the DNA polymerases in eukaryotes
- alpha: primase
- betta: base excision repair and meiotic recombination
- gamma: mitochondrial replication
- delta: nuclear replication, NER & MMR
- epsilon: nuclear replication, NER and MMR
what are telomeres
the ends of eukaryotic chromosomes; allow linear molecules to be replicated w/o loss of information. contain hundreds of tandem repeated copies of a GT rich seq.
How are telomeres synthesized?
Through enzyme telomerase which is a specialized reverse transcriptase bc using an RNA template to make a DNA copy. It has an RNA component that serves as a template for the synthesis of a repetitive GT rich DNA seq ch. of each organism.
Priming can occur in the ____ not the coding DNA
telomere
what are functions of telomeres.
- SEal chromosomal ends
- Attach chromosomes to nuclear envelope
- facilitate replication
- Serve as mitotic clock:
- shorter telomeres induce senescence/apoptosis
- no detectable telomerase act in diff cells.
- telomerase is active in germs cells, stem cells and tumors
- telomeres in fibroblasts from elderly patients are shorter than those from children.
