Proteins Work Together Proteins Work Together in DNA Replication & in DNA Replication &

RepairRepair

Franklin Stahl

The Basic Principle: The Basic Principle: BaseBase Pairing to a Template Pairing to a Template

StrandStrandWatson & Crick’s hypothesis = a Semi-Conservative model

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Original DNASegment

_____?______that hold the 2 strands together are temporarily broken

Hydrogen bonds

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Each original strand serves as a template for a new, complementary strand

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The two new segments are identical to each other AND identical to the original.

Half of each new molecule is comprised of a original template strand (dark blue)

Quick ThinkQuick Think

How DOES DNA’s structure allow for it’s exact replication???

Although Watson & Crick Proposed a Semi-Conservative Model, Other Alternatives Were Proposed

Which Model Does DNA Which Model Does DNA Replication Actually Follow? Replication Actually Follow?

The Meselson-Stahl Experiment:

Quick ThinkQuick ThinkDescribe what semiconservative replication means

DNA Replication: DNA Replication: StartingStartingDNA replication begins at a special sequence of nucleotides which signifies an origin of replication-> proteins pull the two strands apart to form a replication “bubble.” Replication proceeds in both directions:

DNA Replication: DNA Replication: Elongating the Elongating the New StrandNew Strand•DNA polymerases

catalyze the addition of a nucleoside triphosphate to the 3’ end of a growing DNA strand•A nucleoside triphosphate is a nucleotide monomer with 3 P groups the 3 P groups create enough instability that 2 P are lost during the addition of the monomer.•Hydrolysis of the 2 P molecule, pyrophosphate, releases energy that drives the polymerization reaction

DNA Replication: DNA Replication: Antiparallel Antiparallel ElongationElongation•The 2 strands of a DNA molecule are antiparallel to each other; they’re oriented in opposite directions

•DNA polymerases add nucleotides only to the 3’ end of a growing chain. There-fore growth proceeds in the 5’3’ direction

The leading strand is made continuously & in one piece

The lagging strand is made small chunks, Okazaki fragments, in order to follow the 5’3’ rule

Okazaki fragmentsare then joinedtogether by DNA ligase

Leading vs. Lagging Leading vs. Lagging StrandStrand

http://www3.interscience.wiley.com:8100/legacy/college/boyer/0471661791/animations/replication/replication.swf

Priming DNA SynthesisPriming DNA Synthesis•DNA replication cannot begin without a primer

•A primer is a short segment of RNA that has an available 3’ end

•Primase is a special enzyme that constructs the primer sequence from scratch•The leading strand only needs one primer. However the lagging strand needs several primers•DNA pol I replaces RNA primer nucleotides with DNA after each fragment is made•Ligase then fuses fragments

Quick ThinkQuick ThinkDescribe the role of primase, DNA pol I, DNA pol III, and ligase in the formation of the lagging strand.

Summary of Bacterial DNA Summary of Bacterial DNA ReplicationReplication

Helicase unwinds the helix at the replication forkSingle-strand binding protein stabilizes single stranded DNA until it can be used as a templateTopoisomerase keeps DNA from overwinding by breaking & rejoining the DNA ahead of the replication fork

Misconceptions & the Misconceptions & the Replication MachineReplication Machine

Simplified models make it seem like the DNA molecule is stationary and that the various replication enzymes move down the molecule.Contrary to this idea, there is evidence that supports the reverse notion: Replication enzymes form a complex, a “machine,” and the DNA moves through the machine. It’s analogous to fabric moving through a sewing machine.

Proof Reading & Repairing Proof Reading & Repairing DNADNA

•DNA polymerases are the molecules that “proof-read” each nucleotide to ensure that it is complementary to the template•If a mistake is found, the nucleotide is removed

& replaced•If DNA polymerase doesn’t catch a mistake, or if a mismatch occurs after DNA replication, then excision repair is one way to correct the error:

•A nuclease is a special enzyme that cuts DNA during repair

Quick ThinkQuick Think•What are some ways in which DNA may become damaged?

•Why is it important for DNA to be able to repair itself?

Replicating the Ends of DNA Replicating the Ends of DNA MoleculesMoleculesIn linear DNA, such as eukaryotic DNA, there comes a point toward the end of the DNA molecule where a 3’ end is not available, and, therefore, DNA pol III can no longer add nucleotides:

With each replication, the new DNA molecule is becoming shorter & shorter!!!

To mitigate the effects of the progressive shortening, the ends of a DNA molecule are composed of telomeresrepeating sequences of DNA (in humans, TTAGGG) that don’t code for any genes.

Bright yellow=telomeres

The presence of telomeres postpones the erosion of the end of a DNA molecule.Cells that undergo many divisions, like germ cells which produce gametes, contain an enzyme called telomerase. Telomerase catalyzes the lengthening of telomeres so that a zygote will contain cells that have maximum telomere length

Quick ThinkQuick Think

How are telomeres important in preserving eukaryotic genes?

Why does prokaryotic DNA lack telomeres?