BC 14 DNA Replication

Question 1

origins of replication

Answer 1

AT base

multiple origins for euk

specific initiator proteins called Origin binding proteins recognize origin sequences and separate DNA strands locally creating rep bubble

Question 2

DNA Helicases

Answer 2

catalyze separation of dsDNA
moves 5’-3’
requires energy hydrolysis of ATP

Question 3

SSB protein

Answer 3

binds to ssDNA generated by helicases

stabilizes ssDNA

bind cooperatively

NOT enzymes

protect from degradation

Question 4

direction of replication

Answer 4

ALWAYS 5-3

Question 5

leading strand

Answer 5

synthesizes in the 5-3 direction continuously towards replication fork

Question 6

lagging strand

Answer 6

strand being synthesized in the direction AWAY from the replication fork

Okazaki Fragments

Question 7

Key Components of Replication

Answer 7

dNTP

single stranded template

free OH group

RNA primer

Question 8

Elongation

Answer 8

happens fast, does not need to recognize each base, just recognize if they form A:t G:C
-not yet covalently bound

• if H bonding between two correct DNA polymerase catalyzes formation of covalent bond
• FREE OH GROUP CRITICAL
• only proceeds 5-3 where OH is available

Question 9

Beginning of DNA elongation

Answer 9

RNA primer, synthesized by Primase

primase synthesizes short strand of RNA complementary to the DNA template called RNA primer
RNA Primer: short double stranded region consisting of RNA paired to DNA with free OH 3’ group allowing DNA pol to take over

RNA is later removed and gap is filled

Question 10

polymerases

Answer 10

Delta: elongates leading strands and okazaki frags

alpha(contains primase and initiates DNA synthesis) and epsilon (repair): are also involved.

Question 11

Elongation start details

Answer 11

primase associated with DNA pol alpha produces RNA primer and then pol alpha adds about 20 bps to this RNA and dissociates from the template due to low processivity

DNA pol delta adds bp to this RNA DNA primer beginning strand

Question 12

strand completion

Answer 12

RNAase H recognizes RNA DNA duplexes and removes RNA primer

DNA pol refills holes

DNA ligase joins fragments together

Question 13

RNAase H clinical note

Answer 13

HIV virus converts an RNA genome to dsDNA

RNA DNA complex intermediate

Target RNAas H activity of this ezyme is a target for drug development to block HIV genome replication

Didanosine

anti HIV medicine called a Nucleoside reverse transcriptase inhibitor (NRTI). blocks viral RT.
-ddl does not have a hydroxyl group on the 3’ carbon terminating the extension

Question 14

Topoisomerases

Answer 14

helicase action causes the DNA ahead fo the replication fork to twise forming supercoils inhibiting replication.

topoisomerases relive strain by causing nicks int eh DNA that allow one of both strands to rotate relative to the other

I and II

Question 15

topoisomerase I

Answer 15

covalently attaches to a phosphate of the DNA backbone, creating nick in only one strand

other strand rotates

spontaneous reformation of bond seals nick, no ATP required

Question 16

topoisomerase II

Answer 16

ATP hydrolysis to create a double strand break

allows strands to pass eachother faster than unwind

Question 17

Telomeres

Answer 17

end of euk chromosome

TTAGGG, double and single stranded

DNA pol cannot replicate the extreme ends of each strand

no room for new RNA primer

Question 18

telemorase

Answer 18

RNA component that uses telomeric DNA sequences to extend the existing strand of parental DNA

• RNA dependant DNA polymerase. ie uses own RNA template to synthesize DNA
• extends from 3’OH

AAUCCC

Question 19

aging

Answer 19

loss of about 5 bases per cell division

telemorase activity decreases with age

somatic cells lack telomerase activity

biomarkers to annotate age

cancer cells express elevated telomerase