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51 Cards in this Set
- Front
- Back
T/F: The surprising intracy of DNA replication compared to the chemically similar transcription process arises from the need for extreme accuracy in DNA replication so as to preserve the integrity of the genome from generation to generation. |
True |
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T/F: Nearly all known DNA polymerases can only add a nucleotide donated by a nucleoside triphosphate to the free 3'-OH group of a base paired polynucleotide so that DNA chains are extended only in the 5'->3' direction. |
True |
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What is occasionally referred to as DNA's tertiary structure? |
Supercoiling |
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What is the equation for supercoiling? |
L = T + W |
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What is the linking number? |
The number of times that one DNA strand winds about the other; cannot change so long as the two strands are covalently attached in a circle |
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What is the twist? |
Technically, the number of revolutions that one polynucleotide makes about the duplex axis; literally, the number of base pairs divided by 10.5 |
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What is the writhing number? |
The number of times the DNA helix crosses itself |
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T/F: One single-strand nick by an endonuclease is enough to relax a supercoiled DNA. |
True |
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T/F: Gel electrophoresis separates similar molecules on the basis of their compactness, so that the rate of migration of a circular duplex DNA increases with its degree of superhelicity; the agarose gel electrophoresis pattern of population of chemically identical DNA molecules with different linking numbers therefore consists of a series of discreet bands. |
True |
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DNA in physiological solution has _ bp per turn. |
10.5 |
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The supercoiling of DNA is controlled by a remarkable group of enzymes known as... |
DNA topoisomerases (so named because they alter the topological state (linking #) of circular DNA but not its covalent structure. |
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How do Type I and II topoisomerases differ from each other? |
Type I makes transient single-stranded breaks in DNA, while Type II makes double stranded breaks in DNA with the accompanying hydrolysis of ATP --> ADP + P(i) |
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Type _ topoisomerases catalyze the relaxation of supercoils in DNA by changing their linking number in increments of one turn until the supercoil is entirely released. |
I |
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Type IA topos only relax only... |
Negatively supercoiled DNA (it is a nicking-closing enzyme) |
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When type IA topo is heated to denature it in the process of relaxing DNA, what intermediate is found? |
A topo-DNA (phosphotyrosine) intermediate |
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What mechanism does type IA topo operate by? |
A strand passage mechanism |
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What is the mechanism by which type IB topo's operate? |
A "nick and rotate mechanism" |
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T/F: Type IB topoisomerases are said to mediate a controlled rotation mechanism in relaxing supercoiled DNA; this unwinding is driven by the superhelical tension in the DNA and hence requires no other energy input. |
True |
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Why is DNA gyrase unique among topoisomerases? |
Because it generates negative supercoils in DNA |
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T/F: Naturally occurring DNA's negatice supercoiling promotes DNA unwinding, i.e. it's favorable. |
True |
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In what direction is new DNA ALWAYS replicated? |
5' --> 3' |
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Which strand in DNA replication goes toward the direction of the replication fork, and which goes away? |
The leading strand goes toward the replication fork, and the lagging strand goes away from it in Okazaki fragments |
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What are the two enzymes that can catalyze the formation of RNA primers called? |
RNA polymerase, and primase |
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T/F: There are short RNA segments at the beginning (5' end) of each Okazaki fragment. |
True |
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Name the general process that DNA Poly I follows in DNA replication. |
There is nucleophilic attack of the grown DNA chain's 3'-OH group onto the incoming nucleoside triphosphate, which eliminates PP(i) which is then hydrolyzed by pyrophosphatase to keep the reaction going. |
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How does DNA Poly I "choose" the right base pairs to match with the template strand? |
DNA Poly I actually base pairs with the template strand, rather than directly recognizing the incoming base. |
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T/F: Perhaps the simultaneous excision and polymerization activities of Pol I's 5' --> 3' activity protect DNA from the action of cellular nucleases that would further damage the otherwise gapped DNA. |
True |
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What is the second and main function of the 5' --> 3' exonuclease function of DNA poly I? |
To remove the RNA primers from the Okazaki fragments |
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What is nick translation? |
When there is a single-stranded nick in dsDNA, the 5' --> 3' exonuclease function of DNA polymerase I moves the nick towards the 3' end of the DNA, adding new base pairs along the way. (This is often utilized to yield radioactive DNA by placing only radioactive deoxynucleoside triphosphates in the solution to be incorporated into the nick translation) |
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T/F: Pol I's 5'-->3' exonuclease also removes the RNA primers at the 5' ends of newly synthesized DNA while its DNA polymerase activity fills in the resulting gaps. |
True |
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What enzyme acts as a participant in repairing DNA damage via the SOS reponse? |
DNA Polymerase II |
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LECTURE: When determining the way that DNA is replicated by placing it first in dense N15 and then allowing it to replicate in lighter N14 and separating the DNA by density-gradient centrifugation, what results ultimately demonstrated that DNA is replicated semi-conservatively? |
After 1 generation, all strands were of an intermediate weight (1st gen had half heavy parent strand, half light daughter strand); after 2, some were of a very light weight (all new daughter light strands) and some were of the intermediate weight (still half parent and half daughter) and finally with more and more generations the DNA was at the light weight (more new daughter strands were forming with time). |
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What would the density-gradient centrifugation have looked like if DNA replication were conservative? Dispersive? |
If conservative, the dense N15 parent strand would have never gone away; if dispersive, there would be a slew of bands of varying weights all across the plate |
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T/F: DNA Polymerases have a "right hand" structure. |
True |
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T/F: During the addition of a deoxynucleoside triphosphate to the 3' OH end of the daughter strand, one metal ion activates the primer's 3'-OH group for in-line nucleophilic attack on the incoming dNTP's alpha-phosphate group; and another metal ion acts to orient and electrically stabilize the negatively charged triphosphate group. |
Trueq |
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Which function of which DNA polymerase excises mispaired nucleotides from the 3' end of the growing DNA srand? |
The 3' --> 5' exonuclease function of DNA poly I |
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Which enzyme synthesizes a 3' primer for replication of DNA in Okazaki fragments? |
Primase |
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What activity is used to remove RNA primers from the lagging strand Okazaki fragments? |
Nick translation |
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Which enzyme ends nick translation by joining the 5' phosphate to the 3' OH of adjacent nucleotides on a DNA template? |
DNA ligase |
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What are the 3 steps by which DNA Ligase uses NAD+ to catalyze the formation of a phosophodiester bond between the two open ends of a DNA nick? |
1. An adenyl group of NAD+ is covalently transferred to a Lys residue of ligase 2. The adenyl group is then transferred to the 5' phosphoryl terminus of the nick 3. The enzyme (ligase) catalyzes the formation of a phosphodiester bond on the open ends of the nick when the 3' OH group attacks the 5' phosphoryl group and closes the nick |
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What is polymerase processivity dependent on? |
The presence of a B-subunit that is linked to the Poly III holoenzyme complex noncovalently but highly irreversibly |
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What is the actual function of the Beta subunit? |
It serves as a molecular clamp to hold the polymerase onto the replication fork |
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Since the beta subunit acts as a clamp and surrounds the entirety of the helix, it needs a machine to load it onto the helix. What does this? |
A clamp loader; or the lambda complex |
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How is helicase activity different from topoisomerase activity? |
Helicase breaks the H-bonds between base pairs, and topoisomerase breaks the phosphate backbone |
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What proteins in the Poly III holoenzyme prevent the DNA from rehybridizing after being broken apart by Helicase? |
SSB proteins |
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When two forms of DNA are topologically equivalent, it means they are... |
Interconvertible without breaking any bonds |
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Breaking or nicking the phosphate backbone of one DNA strand... |
Releases all of the supercoiling |
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What does the gel pattern of negatively supercoiled DNA treated with Topoisomerase I ultimately indicate? |
That there is a favorable release in Energy, requiring no ATP, when Topo I releases the negative supercoils (think of the rubberband analogy) |
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How does the Topo I enzyme act? Why? |
As a hinge and clamp, which allows the passage of the complementary strand through the single-stranded nick, increasing the Linking number until w = 0 and there is no more supercoiling |
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Which Topo alters W by 2 and increases negative supercoiling? |
Topo II |
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Describe the termination of the replication of a bacterial chromosome. What protein is required for the termination of replication? |
There are 6 termination sites; A, D, and E are right and opposite of oriC and only allow clockwise replication through it; B, C, and F are left and opposite oriC and only allow counter-clockwise replication. The Tus protein is required and binds to these sites and blocks the action of DnaB helicase. |