CHAPTER 16 MOLECULAR BASIS OF INHERITANCE
DNA as genetic material? Deducted that DNA is the genetic material Initially worked by studying bacteria & the viruses that infected them 1928 Frederick Griffiths studied bacterial transformation using Streptococcus pneumonia
Used a pathogenic (disease causing) strain & nonpathogenic strain Killed pathogenic strain w/ heat & mixed with living nonpathogenic strain Some of the nonpathogenic transformed to pathogenic Oswald-Avery deducted that the transforming agent was DNA
Hershey Chase Experiment Used viruses that infect bacteria bacteriophage or phage T2 is common virus of E. coli bacteria of mammal intestines Radioactively tagged sulfur in proteins & the phosphorous in DNA of the T2 phage Infected E.Coli with T2 phage Radioactive sulfur in supernate deducted that protein did not enter bacteria Radioactive phosphorous in cells deducted that DNA enter cells
T2 Phage
Review of DNA Structure
Erwin Chargaff studied the nitrogen bases in the DNA of many species Noticed that amount of adenine equals thymine and amount of cytosine equals guanine Chargaff s rule
Building a Structural Model of DNA Rosalind Franklin & Maurice Wilkins used X - ray crystallography to uncover the double helix of DNA Sugar phosphate backbone on outside Watson & Crick purine paired with a pyrimidine (purine to purine would make helix too wide) Side groups on each nitrogen base forms a hydrogen bond between them
Watson & Cricks Hypothesis of DNA Replication Each strand serves as a template for the new strand
Origin of DNA Replication 1. Replication begins at specific sites where the two parental strands separate & form replication bubbles 2. Bubbles expand laterally, as DNA replication proceeds in both directions 3. Replication bubbles fuse & synthesis of daughter strands is complete
Antiparallel Elongation Enzyme DNA polymerase adds new nucleotides to the template strands Two strands of DNA run antiparallel (opposite directions) DNA polyermase III adds nucleotides in the 5 to 3 direction
Leading strand new strand made in 5 to 3 direction Lagging strand strand produce by nucleotides being added in the direction away from the replication fork ; synthesis occurs with a series of segments called Okazaki fragments DNA Ligase enzyme that joins the sugar phosphate backbones of Okazaki fragments to form new DNA strand
Priming DNA Synthesis DNA polymerases cannot initiate synthesis of a polynucleotide (can only add nucleotides to the 3 end) Primer initial nucleotide chain; short & consist of either DNA or RNA ; initiate DNA replication Primase enzyme that can start a stretch of RNA from scratch
1. Primase joins RNA nucleotide to primer 2. DNA polymerase III adds DNA nucleotides to 3 end of primer 3. Continues adding DNA nucleotides 4. DNA polymerase I replaces the RNA nucleotide with DNA versions
Proteins For Leading & Lagging Strands PROTEIN FUNCTION Helicase Unwinds double helix at replication fork Single-strand binding protein Topiosomerase Stabilizes single-stranded DNA until used as a template Corrects overwinding ahead of replication fork
Protein Primase DNA pol III DNA pol I DNA Ligase Function for Leading Strand Synthesizes a single RNA primer at 5 end of leading strand Continuously synthesizes the leading strand, adding on to primer Removes primer from 5 end of leading strand & replaces it with DNA Joins the 3 end of DNA that replaces the primer to rest of leading strand Function for Lagging Strand Synthesizes an RNA primer at the 5 end of each Okazaki fragment Elongates each Okazaki fragment, adding on to its primer Removes the primer from the 5 end of each fragment & replaces it with DNA, adding on to 3 end of adjacent fragment Joins Okazaki fragments
Summary of DNA replication 1. Helicase unwinds double helix 2. Single-stranded binding proteins stabilize template strands 3. Leading strand is synthesized continuously in 5-3 direction by DNA pol III 4. Primase begins synthesis fo RNA primer for 5 th Okazaki fragment 5. DNA pol III completes synthesis of fourth fragment; when it reaches RNA primer on third fragment it will break off & move to replication fork, and add DNA nucleotides to 3 end of fifth fragment 6. DNA pol I removes primer from 5 end of second fragment, replacing it with DNA nucleotides that it adds one by one to 3 end of third fragment. Replacement of last RNA nucleotide with DNA leaves the sugar-phosphate backbone with a free 3 end 7. DNA ligase binds 3 end of the second fragment to the 5 end of the first fragment
1 2 3 7 4 5 6
Proofreading & Repairing DNA DNA polymerases proofread nucleotides & immediately replace any incorrect pairing Some mismatched nucleotides evade proofreading or occur after DNA synthesis is complete damaged Mismatch repair cells use special enzymes to fix incorrect nucleotide pairs 130 repairing enzymes identified in humans to date
Nucleotide Excision Repair Thymine dimer type of damage caused by UV radiation Buckling interrupts DNA replication
Replicating Ends of DNA Molecules Since only adds to 3 end, no way to complete 5 ends Even if Okazaki fragment is started with a an RNA primer, it can not be replaced with DNA when removed Results in shorter DNA molecules Problem exists only in eukaryotes due to linear DNA Prokaryote DNA is circular
Telomeres nucleotide sequences at ends of DNA molecules Consist of multiple repetitions of a short sequence TTAGGG human telomere Telomeres prevent the erosion of genes near the ends of DNA molecules Somatic cells of older individuals have shorter telomeres related to aging? In germ cells the erosion of telomeres can lead to loss of essential genes Telomerase enzyme that catalyzes the lengthening of telomeres; restores the original length