Molecular Biology of the Gene Chapter 12

Outline Genetic Material Transformation DNA Structure Watson and Crick Eukaryotic DNA Replication

DNA is the Genetic Material Frederick Griffith investigated virulence of Streptococcus pneumoniae Two forms of the bacteria: S strain (smooth) has a capsule surrounding the cell, causes mice to die when this strain is injected R strain (rough) does not have capsule, mice live when this strain injected Heat-killed S strain, mice live Combine heat-killed S strain plus live R strain, mice die, live S strain isolated from dead mice Concluded that some substance needed to make the capsule passed from the dead S strain to the living R strain Virulence passed from the dead strain to the living strain Transformation

Griffith’s Transformation Experiment

Further research by Avery, Macleod, and McCarty Discovered that DNA is the transforming substance DNA from dead cell was being incorporated into genome of living cells Used enzymes: RNases (breakdown RNA) Proteases (breakdown proteases) DNases (breakdown DNA) Treatment of the transforming substance with DNase prevents transformation

Bacteria and Bacteriophages Viruses consist of a protein coat (capsid) surrounding a nucleic acid core. Bacteriophages are viruses that infect bacteria

Hershey and Chase: Radioactively labeled the DNA core and protein capsid of a phage. Results indicated that DNA, not the protein, enters the host. The DNA of the phage contains genetic information for producing new phages.

Hershey and Chase experiments Determined that DNA is the genetic material Experiment 1 Viruses with DNA labeled with 32P were incubated with E.coli Mixed in a blender to remove virus particles attached to cells Centrifuged so bacteria formed a pellet Results- viral DNA was inside the bacteria

Hershey and Chase experiments Viral proteins in capsids were labeled with 35S and viruses were incubated with E.coli Mixed in blender and centrifuged Results- labeled proteins were washed off with the capsids and were not inside the bacteria

Structure of DNA Determined by Watson and Crick Double helix Composed of monomers called nucleotides Each nucleotide has 3 parts Phosphate group Deoxyribose sugar Nitrogenous base- 4 possible Adenine and guanine- purine bases- double ring structure Cytosine and thymine- pyrimidine bases-single ring structure

Erwin Chargaff made key DNA observations that became known as Chargaff’s rule Purines = Pyrimidines A = T and C = G Rosalind Franklin’s (1920-1958) X-ray diffraction experiments revealed that DNA had the shape of a coiled spring or helix

X-Ray Diffraction of DNA

In 1953, James Watson and Francis Crick deduced that DNA was a double helix They came to their conclusion using Tinkertoy models and the research of Chargaff and Franklin James Watson (1928- ) Francis Crick (1916-2004)

Watson/Crick Model of DNA

Overview of DNA Structure

DNA structure- ladder analogy DNA molecule consists of 2 chains of nucleotides Arranged in a “ladder” configuration Alternating phosphate and sugar groups form sides of ladder Rungs are composed of paired nitrogen bases Complementary base pairing-purine with pyrimidine Adenine pairs with thymine Guanine pairs with cytosine The “ladder” then coils to form a helix

Eukaryotic Replication The two DNA strands are held together by weak hydrogen bonds between complementary base pairs A and T C and G Each chain is a complementary mirror image of the other So either can be used as template to reconstruct the other DNA replication occurs in the nucleus. Occurs during S phase of interphase of the cell cycle DNA replication is semi-conservative.

Eukaryotic Replication DNA replication begins at numerous points along linear chromosome. DNA unwinds and unzips into two strands. Each old strand of DNA serves as a template for a new strand. Complementary base-pairing forms new strand on each old strand. Semiconservative: Each daughter DNA molecule consists of one new chain of nucleotides and one from the parent DNA molecule. The 2 daughter DNA molecules will be identical to the parent molecule.

Enzymes Involved in DNA Replication Before replication begins, the 2 strands of the parent molecule are hydrogen-bonded together. The enzyme DNA helicase unwinds and “unzips” the double-stranded DNA. The enzyme primase puts down a short piece of RNA termed the primer for daughter strand synthesis. RNA primers are removed and replaced with DNA. New DNA nucleotides fit into place along divided strands by complementary base pairing. DNA polymerase- reads along each single strand adding the complementary nucleotide DNA ligase repairs any breaks in the sugar-phosphate backbone. Two daughter DNA molecules have now formed that are identical to the original parent molecule.

Semiconservative Replication of DNA Old strand –lavender New strand- hot pink Daughter DNA helix Daughter DNA helix

Ladder Configuration and DNA Replication

Meselson and Stahl’s DNA Replication Experiment

Replication Errors Genetic variations are the raw material for evolutionary change Mutation: A permanent (but unplanned) change in base-pair sequence Some due to errors in DNA replication Others are due to to DNA damage DNA repair enzymes are usually available to reverse most errors