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Introduction to Mol. Biol.ppt
- 2. 1. DNA Replication 2. DNA Transcription 3. Genomic Technology and Bioinformatics
- 3. Chemistry and Metabolism of Nucleic Acids
- 6. Early scientists thought protein was the cell’s hereditary material because it was more complex than DNA Proteins were composed of 20 different amino acids in long polypeptide chains copyright cmassengale 6
- 7. Adenine must pair with Thymine Guanine must pair with Cytosine The bases form weak hydrogen bonds copyright cmassengale 7 G C T A
- 8. Rosalind Franklin took diffraction x-ray photographs of DNA crystals In the 1950’s, Watson & Crick built the first model of DNA using Franklin’s x-rays copyright cmassengale 8
- 12. Two strands coiled called a double helix Sides made of a pentose sugar Deoxyribose bonded to phosphate (PO4) groups by phosphodiester bonds Center made of nitrogen bases bonded together by weak hydrogen bonds copyright cmassengale 12
- 13. A gene is the sequence of nucleotides within a portion of DNA that codes for a peptide or a functional RNA Sum of all genes = genome
- 14. DNA has to be copied before a cell divides DNA is copied during the S or synthesis phase of interphase New cells will need identical DNA strands copyright cmassengale 14
- 15. S phase during interphase of the cell cycle Nucleus of eukaryotes copyright cmassengale 15 Mitosis -prophase -metaphase -anaphase -telophase G1 G2 S phase interphase DNA replication takes place in the S phase.
- 16. Semiconservative Daughter DNA is a double helix with 1 parent strand and 1 new strand Found that 1 strand serves as the template for new strand
- 17. Each strand of the parent DNA is used as a template to make the new daughter strand DNA replication makes 2 new complete double helices each with 1 old and 1 new strand
- 18. Site where replication begins ◦ 1 in E. coli ◦ 1,000s in human Strands are separated to allow replication machinery contact with the DNA ◦ Many A-T base pairs because easier to break 2 H-bonds that 3 H-bonds Note anti-parallel chains
- 19. Begins at Origins of Replication Two strands open forming Replication Forks (Y-shaped region) New strands grow at the forks copyright cmassengale 19 Replication Fork Parental DNA Molecule 3’ 5’ 3’ 5’
- 20. Before new DNA strands can form, there must be RNA primers present to start the addition of new nucleotides Primase is the enzyme that synthesizes the RNA Primer DNA polymerase can then add the new nucleotides copyright cmassengale 20
- 21. Enzyme Topoisomerase attaches to the 2 forks of the bubble to relieve stress on the DNA molecule as it separates copyright cmassengale 21 Enzyme DNA Enzyme
- 22. An enzyme that catalyzes the addition of a nucleotide to the growing DNA chain Nucleotide enters as a nucleotide tri-PO4 3’–OH of sugar attacks first phosphate of tri- PO4 bond on the 5’ C of the new nucleotide ◦ releasing pyrophosphate (PPi) + energy
- 23. Bidirectional synthesis of the DNA double helix Corrects mistaken base pairings Requires an established polymer (small RNA primer) before addition of more nucleotides Other proteins and enzymes necessary
- 24. DNA polymerase can only ADD nucleotides to a growing polymer Another enzyme, primase, synthesizes a short RNA chain called a primer ◦ DNA/RNA hybrid for this short stretch ◦ Base pairing rules followed (BUT A-U) ◦ Later removed, replaced by DNA and the backbone is sealed (ligated)
- 25. Actually how DNA is synthesized ◦ Simple addition of nucleotides along one strand, as expected Called the leading strand DNA polymerase reads 3’ 5’ along the leading strand from the RNA primer Synthesis proceeds 5’ 3’ with respect to the new daughter strand Remember how the nucleotides are added!!!!! 5’ 3’
- 26. Actually how DNA is synthesized ◦ Other daughter strand is also synthesized 5’3’ because that is only way that DNA can be assembled ◦ However the template is also being read 5’3’ Compensate for this by feeding the DNA strand through the polymerase, and primers and make many short segments that are later joined (ligated) together ◦ Called the lagging strand
- 27. 27 The Leading Strand is synthesized as a single strand from the point of origin toward the opening replication fork RNA Primer DNA Polymerase Nucleotides 3’ 5’ 5’ copyright cmassengale
- 28. 28 The Lagging Strand is synthesized discontinuously against overall direction of replication This strand is made in MANY short segments It is replicated from the replication fork toward the origin RNA Primer Leading Strand DNA Polymerase 5’ 5’ 3’ 3’ Lagging Strand 5’ 5’ 3’ 3’ copyright cmassengale
- 29. Other enzymes needed to excise (remove) the primers ◦ Repair polymerase – replaces RNA with DNA ◦ DNA ligase – seals the sugar-phosphate backbone by creating phosphodiester bond Requires Mg2+ and ATP
- 31. Base pairing rules must be maintained ◦ Mistake = genome mutation, may have consequence on daughter cells Only correct pairings fit in the polymerase active site If wrong nucleotide is included ◦ Polymerase uses its proofreading ability to cleave the phosphodiester bond of improper nucleotide Activity 3’ 5’ ◦ And then adds correct nucleotide and proceeds down the chain again in the 5’ 3’ direction
- 32. Helicase opens double helix and helps it uncoil Single-strand binding proteins (SSBP) keep strands separated – large amount of this protein required Sliding clamp ◦ Subunit of polymerase ◦ Helps polymerase slide along strand All are coordinated with one another to produce the growing DNA strand (protein machine)
- 34. Different enzymes recognize, excise different mistakes DNA polymerase synthesizes proper strand DNA ligase joins new fragment with the polymer
- 35. 35 Chemicals & ultraviolet radiation damage the DNA in our body cells Cells must continuously repair DAMAGED DNA Excision repair occurs when any of over 50 repair enzymes remove damaged parts of DNA DNA polymerase and DNA ligase replace and bond the new nucleotides together copyright cmassengale