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Question 1

Primer:
1)
2)

Answer 1

Primer: for the initiation of synthesis

1) a short oglionucleotide chain that is complementary to template strand
2) functions to provide a free 3’OH group for the incoming nucleotide to be added to by a phosphodiester bond
* no known DNA polymerase (DNAP) can initiate synthesis on a bare template. DNAPs always have to have something to add on to

Question 2

dNTPs:

Answer 2

dNTPs: dATP, dCTP, dGTP, dTTP. Monomeric units of DNA molecules. Substrates of synthesis reaction.

Question 3

DNAPs (DNA Polymerases):

Answer 3

DNAPs: enzymes that carry out synthesis

Question 4

DNA synthesis is simply….

Answer 4

DNA synthesis is simply the linkage together of nucleotides (complimentary to template) by a covalent phosphodiester bond

Question 5

Direction of DNA synthesis is always…

Answer 5

Direction of DNA synthesis is always 5’—3’.

Question 6

DNA replication in E. Coli (bacterial system) initiates at the origin called ______.

Answer 6

DNA replication in E. Coli (bacterial system) initiates at the origin called OriC —> chromosomal origin of replication.

Question 7

OriC is a ____ base pair region on the xsomal DNA.

Answer 7

OriC is a 245 base pair region of the xsomal DNA.

Question 8

OriC contains ___________________ that are important in the initiation of DNA synthesis.

Answer 8

OriC contains two sets of repeat sequences that are important in the initiation of DNA synthesis: series of three 13-mer repeats and a series of four 9-mer repeats.
(13-mer is an oglionucleotide 13 bases long)

Question 9

Why are the two sets of repeat sequences in the OriC important?

Answer 9

The repeats are important in binding proteins needed for initiation of synthesis.

Question 10

The OriC repeats are A/T rich…

Answer 10

The repeats are A/T rich, which is important to the function bc A/T pairing represents weak H-bonds and the OriC facilitates the unwinding of the dbl helix.

Question 11

Steps to bacteria DNA replication-initiation:

Answer 11

Initiation: binding of the HU protein and initiation protein to the series 9-mer repeats

• requires ATP for energy to drive bonding rxn
• results in DNA coiling around the proteins. Coiling puts torsional strain on dbl helix.

Question 12

What does initiation result in in bacteria DNA replication?

Answer 12

Initiation results in the DNA coiling around the HU and initiation proteins, and the coiling puts a torsional strain on the dbl helix.

Question 13

-

Answer 13

Unwinding: in order to release the torsional strain caused by the DNA coiling around the HU and I proteins, there is unwinding at the A/T rich regions (weak spot).

• this generates localized region of ssDNA to form a replication bubble
• unwound state is stabilized by binding of a single stranded DNA binding proteins (ssb)

Question 14

What does unwinding generate in bacteria DNA replication?

Answer 14

Unwinding generates a localized region of the ssDNA to form a replication bubble.

Question 15

The unwound state of bacterial DNA is stabilized by:

Answer 15

The unwound state of bacterial DNA is stabilized by: single stranded DNA binding proteins (ssb)

Question 16

Purpose of ssb’s:

Answer 16

Purpose of ssb’s: to keep the helix “open” long enough for DNA to be copied p, by preventing base pairing from reforming the dbl helix

Question 17

The initial unwinding is a physical reaction in response to the _______________.

Answer 17

The initial unwinding is a physical reaction in response to the torsional strain.

Question 18

Further unwinding of the dbl helix is carried out by enzymes called:

Answer 18

Further unwinding of the dbl helix is carried out by enzymes called: helicases. Helicases enzymatically break H-bonds of the dbl helix as replication proceeds from origin.

Question 19

___________ enzymatically break H-bonds of dbl helix as replication proceeds from the origin.

Answer 19

Helicases enzymatically break H-bonds of dbl helix as replication proceeds from the origin.

Question 20

Helicases bind to ssDNA at the _______________ and break H-bonds btwn bases as replication moves forward.

Answer 20

Helicases bind to ssDNA at the replication fork and break H-bonds as replication moves forward.

Question 21

What results from Helicase unwinding?

Answer 21

As a result of helicase unwinding, a superficial knot builds up in front of the replication fork.

Question 22

What works to remove the knot caused by Helicase unwinding the dbl helix?

Answer 22

DNA gyrase is a topoisomerase type 2 that works to remove knots two supercoils at a time.

Question 23

DNA gyrase

Answer 23

DNA gyrase: is a type two topoisomerase that works to remove the knot caused by Helicase two supercoils at a time.

Question 24

DNA gyrase is necessary in order for synthesis to replicate to completion.

Answer 24

DNA gyrase is necessary in order for synthesis to replicate to completion.

Question 25

Steps to bact DNA replication-priming: 1) 2)

Answer 25

Priming: carried out by enzyme primase, a special type of RNA polymerase that functions in DNA replication. 1) primase binds to Helicase and sets down a short oligonucleotide of RNA bases that are complementary to exposed ssDNA template. 2) primer, made by primase, serves to provide a free 3'OH group to which DNAP can add new bases

Question 26

What do all DNAPs require?

Answer 26

All DNAPs require a preexsisting 3'OH group to initiate synthesis. RNAPs do not have this requirement.

Question 27

_______, made by primase, serves to provide a free 3'OH group to which DNAPs can add new bases.

Answer 27

Primer, made by primase, serves to provide a free 3'OH group to which DNAPs can add new bases.

Question 28

All biological DNA synthesis begins w a short ______ primer that needs to be removed before replication is complete.

Answer 28

All biological DNA synthesis begins w a short RNA primer that needs to be removed before replication is complete.

Question 29

Steps to bacteria DNA replication-elongation: - -

Answer 29

Elongation: addition of nucleotides that are complementary to template strand - once DNA is primed, it proceeds in a 5'-3' direction until a terminator signal is encountered or two replication forks meet - elongation is carried out by DNAPs

Question 30

Elongation is carried out by ______.

Answer 30

Elongation is carried out by DNAPs.

Question 31

DNAPs carry out ___________.

Answer 31

DNAPs carry out elongation.

Question 32

``` Characteristics of all DNAPs: 1) 2) 3) 4) 5) ```

Answer 32

1) DNAPs synthesize new strands of DNA that are complimentary and anti parallel to the template 2) DNAPs have, by definition, 5'-3' polymerase activity 3) DNAPs use dNTPs as substrates 4) DNAPs require a primer for initiation 5) DNAPs catalyze the formation of phosphodiester bonds btwn 3'OH of one nucleotide and 5'PO4 of another

Question 33

DNAPs use ______ as substrates.

Answer 33

DNAPs use DNTPs as substrates.

Question 34

DNAPs require a ________ for initiation.

Answer 34

DNAPs require a primer for initiation.

Question 35

3'-5' exonuclease activity:

Answer 35

3'-5' exonuclease activity: used to proofread errors/misincorporated bases

Question 36

5'-3' exonuclease activity:

Answer 36

5'-3' exonuclease activity: used in removal and replacement of RNA primers

Question 37

An exonuclease:

Answer 37

An exonuclease: enzyme activity that works by binding to the end of a DNA molecule and removing/chewing away bases. This removal refers to nuclease: degrades DNA

Question 38

DNAPIII

Answer 38

DNAPIII: main workhorse of replication. It is the one at replication copying all of the DNA. - 5'-3' polymerase activity - 3'-5' exonuclease activity (proofreading)

Question 39

DNAPI

Answer 39

DNAPI: removes the RNA primer with its 5'-3' exonuclease activity and replaces w DNA using its 5'-3' polymerase activity - 5'-3' polymerase activity - 5'-3' exonuclease activity (RNA primer removal)

Question 40

DNAPII

Answer 40

DNAPII: mostly functions in post replication repair - 5'-3' polymerase activity - 3'-5' exonuclease activity (proofreading)

Question 41

``` DNA synthesis is described as being semi discontinuous bc: 1) 2) 3) 4) ```

Answer 41

1) strands of the dbl helix are anti parallel BUT 2) DNA synthesis only,occurs in the 5'-3' direction. 3) therefore, DNA synthesis proceeds in opposite physical directions along the parental template 4) one strand is made continuously (leading strand) and the other is made dis continuously (lagging strand)

Question 42

``` Basic cellular components of replication: 1) 2) 3) 4) 5) ```

Answer 42

1) ssDNA template: what you want to copy 2) dNTPs: monomer of units of DNA molecules. Substrates of synthesis reaction. 3) primer: for initiation of synthesis 4) DNAPs: enzymes that carry out synthesis 5) Mg ions: essential metal cofactor required for the function of DNAPs. Cannot function of lacking Mg ions bc it has no stability