DNA ReplicationDNA Replication

AP Biology Unit 3

(1928) Griffith: Bacteria can be “transformed”/given new traits

Image taken without permission from http://www.nature.com/scitable/topicpage/isolating-hereditary-material-frederick-griffith-oswald-avery-336

(1944) Avery-McCloud-

McCarty: the transforming

substance is DNA

Image taken without permission from http://www.nature.com/scitable/topicpage/isolating-hereditary-material-frederick-griffith-oswald-avery-336

Hershey & Chase Experiment

• Bacteriophages are viruses that infect bacteria

• Bacteriophages consist of DNA and protein (capsid)

• What part of the bacteriophage holds the instructions of how to make more viruses?– DNA

(1952) Hershey & Chase: DNA is the genetic material

Structure of DNA (1953)

• Wilkins and Franklin: Used X-ray crystallography to create an image of DNA; Franklin: sugar & phosphates were the backbone of DNA

• Watson and Crick: Used the X-ray crystallograph to determine the structure of DNA and the base pairing (A-T, C-G)

Image taken without permission from http://www.makingthemodernworld.org.uk/stories/defiant_modernism/01.ST.02/?scene=5

DNA Structure (review)• Double Helix (2 strands of DNA)

• Complementary strands pair up (A & T, C & G)– hydrogen bonds

• Strands are antiparallel (5’ and 3’ ends)

DNA Replication

• When DNA is copied during S Phase of Interphase

• Basic Concept = create a new strand by matching complementary nucleotides to an existing strand

DNA Replication• Replication is semi-conservative (one strand is

old, one strand new)

Origins of Replication

• Where DNA Replication starts

• Differs between organisms– Prokaryotes = 1 origin of

replication– Eukaryotes = many

different origins of replication

General Process of DNA Replication

• Step 1: Initiator Proteins bind to origins of replication to begin replication – Attracts other enzymes involved in replication

process

General Process

• Step 2: – DNA Helicase separates the DNA helix – Topoisomerase (Gyrase) prevents

“overwinding” of DNA by “nicking” the DNA then resealing it.

– Single Stranded Binding Proteins (SSBs) prevent double helix from reforming temporarily

Question…

• What kind of bonds does Helicase break?– Hydrogen bonds

• How do you think SSBs prevent the DNA helix from reforming?– Prevent hydrogen bonds from reforming

General Process

• Step 3: Primase builds an RNA primer at the starting from the 5’ end of the new DNA strand– Uses the 3’ end of existing

DNA strand– Why? Because the DNA

Polymerase III (main DNA building enzyme) needs something to build off of

3’ 5’

5’

Primase

RNA Primer

DNA Polymerase III

General Process

• Step 4: DNA Polymerase III builds the new strand of DNA in a 5’ to 3’ direction – What kinds bonds are

being formed to make a new strand of DNA?

– Covalent bonds

Question…

• What other enzyme builds similarly to DNA Polymerase III?

• RNA Polymerase (don’t mix it up with DNA Polymerase III!)

Problem…

• How can both strands of DNA be replicated in a 5’- 3’ direction at the same time they are antiparallel?

• Answer: leading and lagging strands

Leading and Lagging Strands• Leading strand is

synthesized continuously in the direction of replication (goes in the same direction as helicase)

• Lagging strand is synthesized in short fragments the opposite direction of replication (opposite direction as helicase)

Questions…

• How many primers does the leading strand need?– Only 1– to start replication

• How many primers does the lagging strand need?– Many – one for each Okazaki fragment

Lagging Strand

DNA Ligase

• DNA Ligase seals Okazaki fragments together – Forms covalent bonds between nucleotides to

create a continuous strand of DNA

Finishing DNA Replication

• Problem #1: There are still RNA nucleotides in the DNA (primers)

• Solution = DNA Polymerase I cuts out the RNA nucleotides and replaces them with DNA

Finishing DNA Replication

• Problem #2: Okazaki fragments and bases replaced by DNA Polymerase I are not attached to rest of DNA

• Solution = DNA Ligase seals everything together

Putting It All Together

• Label the diagram on Page 71 with the following terms:- SSBs - DNA Ligase- Leading Strand - Lagging Strand- Helicase - Primase- DNA Polymerase III - Primer- DNA Polymerase I

SSBsDNA Polymerase III

Primase

Primer

DNA Pol III Lagging

Strand

DNA Polymerase I

Leading Strand

DNA Ligase

Helicase