Molecular Genetics

DNA & Replication

DNA Structure

• Nucleotides - Deoxyribose sugar Phosphate Nitrogen Bases

Purine/Pyramidine Pairing

- Adenine/Thymine - Guanine/Cytosine (Chargaff ’ s 1:1 Rule) • Directionality – – 3′ carbon - exposed hydroxyl group 5′ carbon – exposed phosphate group

Chromosome Structure

Euchromatin vs. Heterochromatin

Transposons

• DNA segments that can move to new locations • Can encode transposon enzymes • Effect of insertion mutation: may or may not affect gene expression • In bacteria, can carry antibiotic resistance and jump between genome and plasmids • In humans, some Alu insertions linked to disease (breast cancer, hemophilia, type II diabetes) • Alu insertion in ACE gene linked to athletic performance

Chromosome Structure

• http://www.youtube.com/watch?v=qYsW0jI FH5A (1:17) • http://www.youtube.com/watch?v=eYrQ0E hVCYA (2:18) • http://www.youtube.com/watch?v=gbSIBh FwQ4s (1:42)

DNA Replication

• Semiconservative • ½ old – template strand •½ new – complementary strand • Requires enzymes • Copy: Antiparallel 5′ → 3′ direction

Leading Strand Lagging Strand

Nucleotide Triphosphates supplying energy for bonding

Enzymes/Proteins

• Helicase – Unzips DNA (break H + bonds) to form Replication Fork • SSBPs (single-strand DNA-binding proteins) – Maintains strand separation • Topoisomerases – Breaks/rejoins double helix to prevent knots • Primase – Initiates replication by synthesizing an RNA primer – Inserts at special “ origin of replication ” sequences • DNA Polymerase – Assembles new DNA strand – Unidirectional movement (3′ → 5′ ) – Replaces RNA primer afterwards • DNA Ligase – Connects Okazaki fragments (short segments on lagging strand)

Replication Fork

Okazaki fragments

DNA Replication Animations

• http://www.youtube.com/watch?v= mtLXpgjHL0 (2:04) • http://www.youtube.com/watch?v=I9ArIJW YZHI (2:18)

Telomeres

• The ends of eukaryotic (linear) chromosomes – Repeating nucleotides (vertebrates: TTAGGG x 1000+) • Lagging strand has no primer at one end, leaving a single stranded section of template that will be degraded (shortening the chromosome) • Telomerase adds more repeating bases to the lagging strand – Prevents the lagging strand from getting shorter with each replication –

Absent in somatic cells: chromosomes get progressively shorter over time, limiting lifespan of tissue

– Secret to immortality?

– Cancer cells express telomerase

Telomerase Animations

• http://www.youtube.com/watch?v=AJNoT mWsE0s (2:06) • http://www.youtube.com/watch?v=DV3Xjq W_xgU (4:38) *if time

•

Repairing Mistakes

DNA Polymerase proofreads and

•

Mismatch Repair Enzymes

–

corrects mismatched bases (1/100K Redo section missed by DNA Polymerase times)

• –

Acts as an exonuclease (cleaves Nucleotide Excision Repair System bonds), reducing the error rate to about 1



10 –7

–

Fix DNA damaged by mutagens (ex: UV rays)

Nucleotide Excision Repair

• http://www.youtube.com/watch?v=uN82GL QYAUQ (1:05) • http://www.youtube.com/watch?v=CcTayx Eblio (0:42) • http://www.youtube.com/watch?v=bgUH9 NfO2QM (0:27)