Nucleic Acids Set 4: Dna Replication

Front 1

In what form are bases added to a growing DNA molecule? what is the leaving group?

Back 1

dNTPs. A PPi (pyrophosphate group leaves). This provides the Energy needed for replication.

Front 2

What functional groups are joined?

Back 2

the 3' hydroxyl of one DNA molecule is bound to the phosphate on the 5' Hydroxyl in the adjacent base.

Front 3

How many origins of rep in proks?

Back 3

1

Front 4

How many origins of rep in euks?

Back 4

many

Front 5

are proks haploid, diploid, or triploid?

Back 5

Haploid. (only one copy of chromosome)

Front 6

What are okazaki fragments?

Back 6

short stretches of 5'-->3' DNa that get synthesized on the lagging strand. They are synthesized in the opposite direction of overall DNA growth fork movement.

Front 7

What is the difference between replication in PCr and the cell wrt primers?

Back 7

Pcr primers are DNA, and cell DNA synth primers are RNA.

Front 8

Why is Mg2+ needed for DNA polymerase to work?

Back 8

Mg2+ helps stabilize the charge of the (now) O- on the 3' Carbon (it becomes deprotonated during DNA replication, used to be the 3' OH), this is a better nucleophile to attack the alpha- phosphate than OH. the enzyme + Mg2+ actually lowers the "functional pKa" of this alcohol group, as it is usually around 14-16. Mg2+ interacts with the 2 Aspartate groups within the active site of DNA pol.

Front 9

What is the weakest base pair bond?

Back 9

A=U

Front 10

What Enzyme adds RNA primers on DNA?

Back 10

an RNA polymerase called "Primase"

Front 11

What are the 3 kinds of polymerases we know about (not numbers, or greek symbols)

Back 11

RNA poly (DNA-->RNA), DNA POly (DNA-->DNA) Reverse Transcriptase (RNA--> cDNA)

Front 12

What's a difference between RNA poly and DNA poly, other than their substrates?

Back 12

RNA polys don't have a proofreading function

Front 13

What about DNA POlys STRUCTURE prevents incorrect base pairing?

Back 13

Incorrect base pairing causes steric interference within the active site of DNA poly.

Front 14

What is the error rate in DNA poly w/o proof reading?

Back 14

10^4-10^5

Front 15

What is the error rate in DNA poly w/ proof reading?

Back 15

10^9 - 10^10

Front 16

What are DNA pol I's activities?

Back 16

3'-->5' exonuclease and 5'-->3 exonuclease. It also has polymerase activity, but it's not very efficient, very slow polymerase activity.

Front 17

Which Enzyme (polymerase) has proof reading ability.

Back 17

DNA Pol I (the 3'-->5' exonuclease activity has a proof reading function)

Front 18

Can Poly I go forward if there is a mismatch pair (C-A)

Back 18

No, the -OH group is misoriented. So the DNA pol I goes back (3'-5 exonuclease) to fix the mis-paired base

Front 19

How many ATP equivs does it cost to go back and fix a mis-paired base (1 nucleotide back)

Back 19

2 ATP equivs.

Front 20

What activity does does the larger, 70 kDA polymerase I fragment have (known as the Klenow Fragment)

Back 20

DNA polymerase and 3'-->5' exonuclease

Front 21

What activity does the smaller, 30 kDA polymerase I fragment have?

Back 21

5'-->3' exonuclease

Front 22

What direction of exonuclease function is important in nick translation

Back 22

5'-->3' exonuclease.

Front 23

Where is nick translation common?

Back 23

at okazaki fragments.

Front 24

What is at the 5' end of a nick in DNA?

Back 24

3 phosphate groups (2 get removed when joining the phosphate group to the 3' hydroxyl group

Front 25

What is the 5'-->3' exonuclease activity of polymerase used for primarily in DNA synthesis?

Back 25

Removing the RNA primers.

Front 26

What functional group does DNA ligase have in it's active site?

Back 26

Amino group (NH3+)

Front 27

How many ATP equivalents does ligation cost?

Back 27

2 ATP equivs

Front 28

What initially binds to the DNA ligase in euks?

Back 28

AMP binds to the NH3 group

Front 29

WHat group transfers the AMP group onto DNA ligase in EUKS? what leaves?

Back 29

ATP, and PPi leaves

Front 30

WHat transfers the AMP group onto DNA ligase in proks? What leaves?

Back 30

NAD+ is transferred on, and NMN leaves.

Front 31

Where does AMP from DNA ligase get transferred onto?

Back 31

the free 5' phosphate group on the DNA molecule at the nick site

Front 32

What is the leaving group when the 3'-hydroxyl attacks the 5' phosphate group in nick closure (ligation), catalyzed by DNA ligase?

Back 32

AMP

Front 33

Is DNA pol III in proks or Euks? what does it do?

Back 33

Proks. It is involved in both leading and lagging strand synthesis.

Front 34

What is another name for the processivity factor? what does it do?

Back 34

the B-Clamp. It ensures that DNA pol III finishes the job.

Front 35

What do ssDNA binding proteins do in DNA replication (ssDBP)

Back 35

They bind to the single stranded DNA to prevent the strands from coming back together.

Front 36

What does helicase do?

Back 36

Unwinds the DNA using ATP.

Front 37

What is the main enzyme required for synthesizing DNA (just making the leading and the lagging strand?)

Back 37

DNA pol III

Front 38

What are the 3 phases of DNA replication?

Back 38

Initiation, elongation, termination.

Front 39

What sequence in the OriC region of proks rich in?

Back 39

A/T residues- easy to pull apart.

Front 40

HOw does the OriC region get unwound in prokes?

Back 40

8 DnaA proteins bound to ATP (DnaA-ATP) bind to the DNA to form a right hand solenoid, which put pressure on the OriC site, which causes the DNA to separate. DnA-ADP is removed from the DNA after. This requires ATP.

Front 41

DO both plasmids and chromosomes require a termination (ter) site, and an ori site?

Back 41

Yes

Front 42

What protein binds the ter site in proks?

Back 42

tus

Front 43

What isomerase is used to separate the catenanes during DNA replication?

Back 43

Topoisomerase IV

Front 44

Is the replication fork Faster or slower in Euks?

Back 44

Slower in Euks

Front 45

What is the Rate limiting step in DNA synth in Euks?

Back 45

Removal of the histone protein.

Front 46

What removes the primers in EUK DNA rep?

Back 46

RNase H and MF 1 (5'-->3' exonuclease)

Front 47

WHat is the replication substrate in Euks?

Back 47

Chromatin

Front 48

What protein is bound to telomeres?

Back 48

The Telomere duplex DNA binding protein

Front 49

What type of overhang occurs at the ends of chromosomes (telomeres) after euk DNA replication?

Back 49

3' overhang.

Front 50

What enzyme adds telomeres to the Chromosome, and what does it contain?

Back 50

Telomerase: Protein + RNA (template)

Front 51

What polymerase is Telomerase similar to? and why?

Back 51

Reverse transcriptase because it uses an RNA template (w/i the telomerase protein) to make DNA