Chapter 16 Molecular Basis of. Chapter 16. Inheritance (DNA structure and Replication) Helicase Enzyme

Transcription

1 Chapter 16 Chapter 16 Molecular Basis of Inheritance (DNA structure and Replication) Helicase Enzyme

2 The Amazing Race What is the genetic material? DNA or protein?

3 1928 Griffith transformation of pneumonia bacterium

4 Question generated from Griffith What was the heritable agent, protein or DNA?

5 1944 Avery further studied transformation by destroying lipids, CHO, and proteins

6 Conclusions of Avery, et. al DNA is heritable agent. Results, unfortunately, were not enough to sway the opinion of most scientists. How could one molecule cause so much variation?

7 1947 Chargaff Quantified purines and pyrimidines Suggested base pairing rules (A=T, C=G)

8 1950 Wilkins and Franklin DNA X-rays (a) Rosalind Franklin (b) Franklin s X-ray diffraction photograph of DNA

9 1952 Hershey and Chase bacteriophages incorporation of radioactive viral DNA in new phages

10 EXPERIMENT Batch 1: radioactive sulfur ( 35 S) Phage Bacterial cell Radioactive protein DNA Radioactive DNA Batch 2: radioactive phosphorus ( 32 P)

11 EXPERIMENT Phage Bacterial cell Radioactive protein Empty protein shell Batch 1: radioactive sulfur ( 35 S) DNA Phage DNA Radioactive DNA Batch 2: radioactive phosphorus ( 32 P)

12 EXPERIMENT Phage Bacterial cell Radioactive protein Empty protein shell Radioactivity (phage protein) in liquid Batch 1: radioactive sulfur ( 35 S) DNA Phage DNA Centrifuge Radioactive DNA Pellet (bacterial cells and contents) Batch 2: radioactive phosphorus ( 32 P) Centrifuge Pellet Radioactivity (phage DNA) in pellet

13 Video of Hershey Chase Expt

14 Significance of Hershey & Chase Expt. Unambiguously proved that DNA is the agent of heredity, not proteins.

15 1953 Watson and Crick DNA Model

16 1962 Nobel Prize awarded to Watson and Crick and Wilkins ** Conclusion: Now that we know DNA s structure, we can figure out what it does and how it does it.

17 DNA Replication

18 Models of DNA Replication

19 Semi-Conservative Model (1950s - Meselson and Stahl)

20 Meselson & Stahl Experiment

21 Fun DNA Replication Facts 6 billion bases in human cell = 2 hours of replication time 500 nucleotides added per second Accurate (errors only 1 in 10,000 base pairs)

22 Anti- Parallel Structure of DNA

23 Origins of Replication = Special site(s) on DNA w/specific sequence of nucleotides where replication begins Prokaryotic Cells = 1 site (circular DNA) Eukaryotic Cells = several sites (strands) Mechanism of Replication Step 1

24 Steps 2-5 Helicase: (enzyme) unwinds DNA helix forming a Y shaped replication fork on DNA Replication occurs in two directions, forming a replication bubble To keep strands separate, DNA binding proteins attach to each strand of DNA Topoisomerases: enzymes that work w/helicase to prevent knots during unwinding.

25 Step 6 - Priming Priming = due to physical limitation of DNA Polymerase, which can only add DNA nucleotides to an existing chain RNA primase initiates DNA replication at Origin of Replication by adding short segments of RNA nucleotides. Later these RNA segments are replaced by DNA nucleotides by DNA Pol.

26 Step 7 DNA Pol. = enzyme that elongates new DNA strand by adding proper nucleotides that basepair with parental DNA template DNA Pol. can only add nucleotides to the 3 end of new DNA, so replication occurs from a 5 to 3 direction Leading vs. Lagging Strand results

27 Leading vs. Lagging Strand Leading Strand: strand that can elongate continuously as the replication for progresses Lagging Strand: strand that cannot elongate continuously and moves away from replication fork. Short Okazaki fragments are added from a 5 to 3 direction, as replication fork progresses.

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29 Step 8 DNA Ligase = enzyme that ligates or covalently bonds the Sugar-Phosphate backbone of the short Okazaki fragments together Primers are required prior to EACH Okazaki fragment

30 Overview of DNA Replication DNA i

31 DNA Replication Fork

32 Step 10: Fixing Errors DNA Pol. Proofreads as it elongates Special enzymes fix a mismatch nucleotide pairs Excision Repair: Nuclease: Enzyme that cuts damaged segment DNA Pol. Fills in gap with new nucleotide

33 Mutations Thymine Dimers (covalent bonding btwn Thymine bases) often caused by overexposure to UV rays DNA buckeling skin cancer results, unless corrected by excision repair Substitutions: incorrect pairing of nucleotides Insertions and Deletions: an extra or missing nucleotide causes frameshift mutations (when nucleotides are displaced one position)

34 Problems with Replication Since DNA Polymerase can only add to a 3 end of a growing chain, the gap from the initial 5 end can not be filled Therefore DNA gets shorter and shorter after each round of replication

35 Solution? Bacteria have circular DNA (not a problem) Ends of some eukaryotic chromosomes have telomeres at the ends (repeating nucleotide sequence that do not code for any genes) Telomeres can get shorter w/o compromising genes Telomerase = enzyme that elongates telomeres since telomeres will shorten

36 Telomerases are not in most organisms Many multicellular organisms do not have telomerases that elongate telomeres (humans have them in some cells) So, telomeres = limiting factor in life span of certain tissues Older individuals typically have shorter telomeres

37 Homework Watch Bozeman Videos #27 DNA & RNA Parts 1 and 2 (2 videos)