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43 Cards in this Set

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mutation rate in bacteria
1 nucleotide change per 10^9 nucleotides per cell generation
fibrinopeptides
a 20 amino acid fragment used to determine mutation rates in mammals, their function does not depend on their aa seq., they can tolerate almost any amino acid change
mutation rate in mammals
a protein 400 aa’s long will mutate once every 200,000 years
mutation rates in different proteins
different proteins evolve at very different rates
human genome mutation rate
also 1 nucleotide change per 10^9 nucleotides/division, 3* 10^9 nucleotides in humans, translates to about 3 nucleotides that change in each cell division
DNA duplication rate
1000 nucleotides/second
DNA polymerase
the first nucleotide polymerizing enzyme, discovered in 1957, free nucleotides (deoxyribonucleoside triphosphate) act as the substrates for this enzyme, cannot start a new polynucleotide chain
what underlies DNA replication and repair
DNA base-pairing
directionality of growth for the primer strand (new strand)
5’ – 3’ (OH end)
what drives DNA replication and addition of new nucleotide
breakdown of pyrophosphate to two free phosphates
classification of DNA replication
semiconservative; each original strand acts as a template for a new strand
replication forks
are asymmetric
DNA polymerase and proofreading
first step in proofreading, higher affinity for correct nucleotide, it double checks bec. if it is the wrong nucleotide it dissociates, has a polymerizing site and an editing site
exonucleolytic proofreading
2nd step of proofreading, 3’ – 5’ proofreading found on the DNA polymerase removes wrong nucleotide, exposing a proper 3’ OH end and continuation of replication
DNA primase
uses ribonucleoside triphosophates to synthesize short RNA primers (about 10 nucleotides long) for the start of DNA replication on the lagging strand, DNA polymerase takes over and when it reaches the RNA primer it is replaced with DNA (by RNAse H)
DNA ligase
used to connect the new 3’end to the old 5’ end, joins Okazaki fragments together, uses ATP to activate the 5’ end at the nick before forming the new bond
DNA helicases
hydrolyze ATP when they are bound to single strands of DNA, use this to propel themselves along a single strand of DNA, pry at rates up to 1000 nucleotide pairs/second
single strand DNA binding proteins (SSB proteins)
AKA helix-destabilizing proteins, bind tightly and cooperatively to exposed single-stranded DNA strands without covering the bases and stabilized the unwound conformation, prevent single-stranded hairpin tangles
sliding ring
keeps the DNA polymerase connected to the DNA, releases when it reaches a double strand, requires ATP degradation to bind the halves of the sliding clamp, there is a clamp loader present that loads the clamp and dissociates once the DNA polymerase binds
primosome
found in prokaryotes that link the primase with a DNA helicase, on the lagging strand
strand-directed mismatch repair
detects the potential for distortion in the DNA helix that results from the misfit between non complementary base pairs, only removes the mismatch at the newly synthesized strand, methylates an A at a GATC site, miss the ones that are newly synthesized so can differentiation this strand, eukaryotic cells have nicks on the new strand to differentiate it, three step process of repair:
1. recognition of a mismatch
2. excision of the segment of DNA containing the mismatch
3. resynthesis of the excised segment using the old strand as a template
MutS
part of the strand-directed mismatch repair, binds specifically to a mismatched base pair
MutL
part of the strand-directed mismatch repair, scans the DNA for a nick, once found it triggers the degradation of the DNA all the way back to the mismatch
DNA topoisomerase
prevents DNA tangling during replication, a reversible nuclease
topoisomerase I
produces a transient single-stand break in the backbone, allows for free rotation relieving tension, rapid resealing and does not require additional energy input, forms a covalent bond w/ DNA on only 1 strand
topoisomerase II
forms a covalent bond with both strands where two double helixes cross each over other, makes a double strand break, separates two interlocked DNA circles, uses ATP hydrolysis to:
1. break one double helix reversible to create a DNA gate
2. causes the second, nearby double helix to pass through this break
3. reseals the break and dissociates from the DNA
Mammalian replication fork
1. uses two different DNA polymerases (alpha and delta), alpha starts, pass to delta
2. DNA primase is a subunit of DNA polymerase alpha
spontaneous alterations likely to require DNA repair
1. spontaneous oxidative damage
2. hydrolytic attack
3. uncontrolled methylation by the methyl group donor S-adenosylmethonine
most frequent spontaneous chemical reactions
1. depurination-can release guanine, as well as adenine, from DNA, can lead to deletion of an amino acid pair
2. deamination-converts cytosine to an altered DNA base, uracil, can lead to a substituation of a wrong aa
thymine dimmers
formed from ultraviolet radiation that can produce a covalent linkage between two adjacent pyrimidine bases in DNA
types of genetic recombination
general and site-specific
general recombination
AKA homologous recombination, genetic exchange takes place between a pair of homologous DNA sequences, ex: meiosis crossing over
DNA synapsis
where base pairs form between complementary strands from the two DNA molecules, only occurs when DNA molecules contain long regions of match (or nearly matching) sequences
endonuclease and exonuclease in DNA recombination
endonuclease cuts both strands of the double helix
exonuclease chews the 5’ end creating protruding single-stranded 3’ ends, these free ends find a homologous pair leading to the formation of a joint molecule
DNA hybridization
occurs when a rare random collision juxtaposes complementary nucleotide sequences on two matching DNA single strands forming a short strand of double helix (helix nucleation), rapid zippering then occurs and matching completes, trial and error similar to DNA recombination method
RecA protein
found in E. coli, has a central role in the recombination between chromosomes, make synapsis possible, binds to single stranded protein to form a nucleoprotein filament while at the same time holding a double strand, forms a three strand intermediate, catalyzes unidirectional branch migration
branch migration
occurs after DNA synapsis has occurred, enlarges the heterodulplex region, usually spontaneous but not likely to complete recombination because it goes back and forth, RecA is unidirectional (ATPase)
unidirectional RecA branch migration
is a DNA-dependent ATPase, binds to DNA more tightly when ATP is bound
Rad51
RecA homolog in humans, catalyzes a synaptic reaction between a DNA single strand and a DNA double helix, is required to repair replication forks
Holliday junction
involved in general recombination, initially forms a structure containing two crossing strands and two noncrossing strands, then it isomerizes to form an open structure, then further isomerized to interconvert the crossing and noncrossing strands originally
open form of the holliday junction
some proteins can engage with the junction to move the crossover point rapidly along the two helices, extending the region of heteroduplex DNA
resolution of the holliday junction
occurs when the crossing strands are cut, can lead to obtainment of the original strands or exchanged strands
gene conversion
occurs when meiosis yields three copies of the maternal version and one copy of the paternal allele, occurs through general recombination, can occur during meiosis if there is a slight difference in a heteroduplex and mismatch repair occurs deleting one allele and making an extra copy of the other allele, the DNA then replicates and an extra allele of one of the halves is present