BS1030 Topic 3 Lecture 2 The structure of the DNA double helix

Who discovered the structure of DNA?

James Watson and Francis Crick in 1953.

What key statement did Watson and Crick make about DNA?

"We wish to suggest a structure for the salt of deoxyribonucleic acid (DNA). This structure has novel features of considerable biological interest."

What three pieces of evidence led to the discovery of DNA structure?

Chargaff's base composition data, Rosalind Franklin's X-ray diffraction, and Pauling & Corey's incorrect triple-stranded model.

What did Chargaff's rules show?

Purines = Pyrimidines (A = T, C = G) but A/T does not equal C/G overall.

What did Franklin and Wilkins contribute?

X-ray diffraction images revealing DNA's helical structure.

What was wrong with Pauling and Corey's model?

It had three strands with phosphates on the inside—chemically unstable.

Where must the phosphate groups be in DNA?

On the outside of the helix.

What does base pairing mean?

A pairs with T (2 hydrogen bonds), G pairs with C (3 hydrogen bonds), giving equal spacing across the helix.

Why must base pairs be purine-pyrimidine?

To maintain a constant diameter of the double helix.

What is the overall structure of B-form DNA?

A right-handed double helix with antiparallel strands and complementary base pairing.

What are the antiparallel directions of DNA strands?

One runs 5′→3′, the other 3′→5′.

What are the dimensions of B-form DNA?

10 base pairs per turn (34 Å per turn) and 20 Å in diameter.

What does "phosphodiester link" mean?

The covalent linkage between the 3′ hydroxyl of one nucleotide and the 5′ phosphate of the next.

What stabilises DNA base pairing?

Hydrogen bonds and base stacking interactions.

What other DNA forms exist besides B-DNA?

A-form DNA (dehydrated), Z-form DNA (left-handed), and G-quadruplexes.

What is A-form RNA?

A right-handed double helix formed by RNA due to ribose sugar conformation.

What are the major and minor grooves of DNA?

Alternating surface features where proteins can interact with bases.

Where do most proteins bind in DNA?

In the major groove, where base pair edges are more accessible.

How long is the E. coli genome?

~4 million base pairs, about 1.4 mm in length.

How long is the human genome?

~6 billion base pairs across 46 chromosomes, totaling about 2 meters of DNA per cell.

What are circular DNA molecules?

Closed loops of DNA found in bacteria, mitochondria, chloroplasts, and sometimes eukaryotic genomes.

What is DNA supercoiling?

Twisting of circular DNA to compact it and reduce viscosity.

What are topoisomers?

DNA molecules that differ in the degree of supercoiling.

Which enzymes interconvert topoisomers?

Topoisomerases.

What wavelength does DNA absorb UV light at?

260 nm.

What is hyperchromism?

Increased UV absorbance when DNA strands separate during denaturation.

What is DNA denaturation (melting)?

Separation of double-stranded DNA into single strands due to heat or pH changes.

What is renaturation (annealing)?

Rejoining of complementary DNA strands when cooled slowly.

What are the main enzymes that manipulate DNA?

Nucleases, ligases, polymerases, kinases, phosphatases, methylases, demethylases, and topoisomerases.

What do exonucleases do?

Remove nucleotides from the ends of DNA.

What do endonucleases do?

Cut DNA internally—either randomly or at specific sequences.

What are restriction endonucleases?

Sequence-specific enzymes that cut DNA at palindromic sites.

Why do restriction enzymes recognise palindromic sequences?

Because they bind as dimers to symmetrical DNA sequences.

What are sticky ends?

Single-stranded overhangs left by restriction enzyme cleavage.

What are blunt ends?

Ends with no overhangs after DNA cutting.

What can happen to DNA fragments with matching sticky ends?

They can be ligated together to form recombinant DNA.

What enzyme catalyses the joining of DNA fragments?

DNA ligase.

What reaction does DNA ligase perform?

Recreates a phosphodiester bond between a 3′-OH and a 5′-phosphate using ATP.

Who discovered DNA polymerase I?

Arthur Kornberg in 1958.

What is the role of DNA polymerase I?

Adds nucleotides to a primer's 3′-OH end, using a template strand.

How fast does DNA polymerase I work?

Adds about 10 nucleotides per second.

What is proofreading in DNA replication?

The removal and replacement of mismatched nucleotides by DNA polymerase.

What is meant by "processivity" in polymerases?

The number of nucleotides a polymerase adds before dissociating from the template.

What does DNA polymerase require to start synthesis?

A template strand and a primer with a free 3′-OH group.