The Molecular Basis of Inheritance Chapter 16

The search for genetic material began with bacteria Prokaryotic organisms typically have one circular chromosome, which codes for fewer proteins than the typical linear eukaryote chromosome. Plasmid – circular piece of DNA commonly found in bacteria, in addition to their chromosomes. Bacteria can transmit genetic info through the exchange of plasmids. A. Griffith (1928) found that bacteria could undergo “transformation” by assimilating info from another cell.

The Search Continues… Griffith's experiments hinted that protein was not genetic material. B. Avery’s (and McCarty-MacLeod) Experiment (1944): studied effects of enzymes on transformation; discovered that transforming agent was DNA. Most scientists still believed it was protein; complex molecules and little was known about DNA. C. Hershey and Chase (1952): used bacteriophages and radioactive markers; discovered that DNA is the genetic material.

Understanding DNA Structure D. Chargaff (1947): separated nitrogenous bases in DNA of different organisms. Amount and ratios of bases vary from one species to another; source of genetic diversity. Amount of adenine (A) = amount of thymine (T), and the amount of guanine (G) = amount of cytosine (C).

Discovery of DNA Structure E. Watson & Crick / Wilkins & Franklin (1953): discovered the 3-dimensional structure of DNA; subsequently described the mechanism for DNA replication. After seeing an X-ray crystallography photo of DNA produced by Franklin, Watson and Crick proposed: A ladder-like molecule twisted into a spiral, with sugar-phosphate backbones as uprights and pairs of nitrogenous bases as rungs. The two sugar-phosphate backbones of the helix are antiparallel (run in opposite directions). Pairs of bases hydrogen bond; adenine can only pair with thymine, and guanine with cytosine [purine (A, G) with pyrimidine (C, T)].

DNA Replication Watson and Crick proposed a semiconservative model for DNA replication using a template mechanism. When a double helix replicates, each of the two daughter molecules will have one old or conserved strand and one newly created strand. Although DNA replication is a complex process, it is fast and accurate. In prokaryotes, 500 nucleotides can be added per second; It takes only a few hours to copy the 6 billion bases of a single human cell.

Meselson-Stahl Experiment

Steps in Replication 1. Strand Separation Helicase enzymes bind to the double helix to catalyze unwinding and unzipping of the parent strand. As the helix opens, replication forks spread in both directions creating a replication bubble. Other proteins keep the separated strands apart and stabilize the unwound DNA until new complementary strands can be synthesized. 2. New DNA Synthesis DNA polymerase catalyzes the addition of free-floating nucleotides to the DNA strand. According to base-pairing rules, new nucleotides align themselves along the templates of the old DNA strands.

Steps in Replication (cont) Sugar phosphate backbones of DNA strands run in opposite directions (antiparallel). A DNA strand has a 3' end, on which a hydroxyl group is attached to carbon 3 of the terminal deoxyribose. At the 5' end, a phosphate group is attached to carbon 5 of the terminal deoxyribose. DNA polymerase can only make strands in the 5' to 3' direction. Leading strand -- DNA strand which is synthesized as a single polymer in the 5'  3' direction. (continuous synthesis) Lagging strand -- DNA strand is synthesized in an overall 3’  5’ direction. (discontinuous; Okazaki fragments)

Steps in Replication (cont) 3. Sealing the DNA Primer -- Short RNA segment (about 10 nucleotides) that is complementary to a DNA segment and that is necessary to begin DNA replication. Only one primer is necessary for replication of the leading strand, but many primers are required to replicate the lagging strand (each Okazaki fragment). DNA polymerase removes the RNA primer and replaces it with DNA. DNA ligase catalyzes the linkage between each new Okazaki fragment the growing chain. 4. Re-winding the DNA Animation