Structure

Sugar phosphate backbone on the outside of the helix. Bases are on the inside of the stand. The bases from each strand bond with each other.

Chargaff's Rules

Because of DNA base pairing rules and the double helix shape of DNA:

• The number of A equals the number of T
• The number of C equals the number of G

DNA

All DNA contains nitrogen.

Made of a sugar, phosphate, and base.

DNA runs anti-parallel to the other strand.

Base pairing is dictated by hydrogen bonds between the bases.

The Phosphate is negatively charged, making the DNA negatively charged.

Purines

Larger, double ring.

Adenine (A)

Guanine (G)

Pyrimidines

Smaller, single ring.

Cystine (C)

Thymine (T)

Big rings go with small rings (A-T, C-G).

DNA has Directionality

(like a one-way street)

5' carbon has the phosphate group attached (phosphate first).

• Phosphate is denoted by an open circle: ◯.

3' carbon has a hydroxyl (-OH) group attached first.

5'-ATCG-3' has anti-parallel stand of 3'-TAGC-5'.

3'

3 Prime

5'

5 Prime

Replication

Replication Enzymes

Helicase

Enzymes that untwists the double helix dna.

The sites they work at are called replication forks.

Primase

Synthesizes a short RNA primer to serve as the initial nucleotide chain for DNA Polymerase III.

This primer is five to ten nucleotides long.

DNA Polymerase I

Removes all RNA primers.

RNA primers have to be removed and replaced.

Gaps must be filled by enzyme DNA ligase so there are no gaps in the DNA.

Ligase

Joins together the newly created fragments of DNA.

Topoisomerase

Relieves the strain on unwound double strand dna.

Holds onto the double sided dna strands just before the fork to stabilize it.

Origin of Replication

A sequence of dna that enzymes bind to where replication starts.

There are multiple sites of origin to speed up the process. Because the replication process is linear, it is otherwise slow.

Single Stand Binding Proteins

Stabilizes the vulnerable single-strand dna.

DNA Polymerase III

Requires a short rna primer to start from.

Used to actually replicate the dna strand.

Requires a free 3'-OH group? What is this?

The new strand built by DNA polymerase III towards the fork is called the leading strand.

Builds the new strand in the 5' to 3' direction. That's the only direction it can move.

• This is opposite of the parental strand's direction.

• From the parental strand's perspective, DNA pol III builds from 3' to 5'.

RNA Primer

First 3 bases of each strand.

Synthesized by the enzyme primase.

Lagging Strand Okazaki Fragments

Has to fill the gap left from the opposite end fork. Has to build in the 5' to 3' direction. So new fragments are created behind the trailing fork that splits in the opposite direction and these fragments build back to front to connect to the leading strand / last okazaki fragment. Each fragment starts with the 3 nucleotides closest to the opposite fork.

Mismatch Repair System

• Specialized proteins scan newly replicated DNA
• Detect mismatched base pairs (like A-C or G-T)
• Cut out incorrect nucleotide
• DNA polymerase fills in with correct nucleotide
• Ligase seals the gap

Theories of Replication

Semi-Conservative DNA Replication This is the way DNA replicates

The two strands come apart – each acts as a template for synthesis of a new complementary strand.

Conservative DNA Replication

Original parent double strands remain intact and two completely new double helix strands are synthesized.

Dispersive Replication

Both original parent strands are broken up into small pieces and incorporated into newly synthesized strands.

Meselson-Stahl Experiment

Bacteria cultured in flask w/ heavy nitrogen isotope. Then bacteria transferred to a flask w/ a light nitrogen isotope and allowed to replicate. Over generations, 1st generation was hybrid weight, over time, more light dna was created. Density splits because the hybrid stands in the first generation split and form one hybrid dna and another solely light dna. Density splits over each new generation after the first. Dna uses semi-conservative replication.

Heavy-Nitrogen DNA Strands (▮) Replicated in a Solution of Light Nitrogen Isotopes (▯)

         ▮▮
        🡗   🡖
    ▮▯       ▮▯
   🡗  🡓       🡓  🡖
▮▯  ▯▯    ▮▯  ▯▯