The Molecular Basis of Inheritance Chapter 16

DNA Molecule Deoxyribose, a five-carbon sugar Phosphate group Nitrogenous base (A,T,G,C) Anti-parallel One strand is the “sense” strand or the one that holds the gene The other strand is the “template” or anti-sense strand and has the gene’s copy and will be copied into an mRNA

Frederick Griffith - 1928 Griffith's experiment was conducted in 1928 by Frederick Griffith which was one of the first experiments suggesting that bacteria are capable of transferring genetic information, otherwise known as the “transforming principle”, which was later discovered to be DNA.

Griffith Experiment – DNA

Hershey-Chase experiment The was a series of experiments conducted in 1952 by Alfred Hershey and Martha Chase that identified DNA to be the genetic material of phages and, ultimately, of all organisms. Before this experiment was conducted, it was said that proteins were the genetic material in viruses, not DNA.

Chargaff’s Rule: %A=%T and %G=%C Erwin Chargaff (1905–2002) was an Austrian biochemist who emigrated to the United States during the Nazi era. Through careful experimentation, Chargaff discovered two rules that helped lead to the discovery of the double helical structure of DNA. Watson and Crick used this information to discover the DNA structure as we know it. Chargaff had told them about a simple relationship he had found between the occurrence of different bases in DNA: adenine and thymine were present in roughly the same amounts and so were guanine and cytosine. One of each pair was a larger purine; the other, a smaller pyrimidine.

Incorporation of a nucleotide into a DNA strand Phosphodiester Bonds Nucleotide tri-phosphates lose 2 phosphates to release enough energy to for polymerization of nucleotides to form DNA The incoming nucleotide triphosphate is added to the 3’ OH of the preceding sugar

Semi Conservative Replication Read Meselson-Stahl experiment, pages 284, 285

DNA REPLICATION

Bidirectional Replication In Prokaryotes (Circular DNA) - 1 replication fork In Eukaryotes (Linear DNA) - Multiple replication forks

Enzymes Involved in DNA Replication DNA Helicases - These proteins bind to the double stranded DNA and stimulate the separation of the two strands. DNA single-stranded binding proteins - These proteins bind to the DNA as a tetramer and stabilize the single-stranded structure that is generated by the action of the helicases. Replication is 100 times faster when these proteins are attached to the single-stranded DNA. DNA Gyrase (Topoisomerase) - This enzyme catalyzes the formation of negative supercoils that is thought to aid with the unwinding process. In addition to these proteins, several other enzymes are involved in bacterial DNA replication. DNA Polymerases - DNA Polymerase I (Pol I) was the first enzyme discovered with polymerase activity, and it is the best characterized enzyme. It is not the primary enzyme involved with bacterial DNA replication. That enzyme is DNA Polymerase III (Pol III). Three activities are associated with DNA polymerase I; 5' to 3' elongation (polymerase activity) 3' to 5' exonuclease (proof-reading activity) 5' to 3' exonuclease (repair activity) DNA Polymerase III (Pol III) is the enzyme that performs the 5'-3' repair function. RNA Primase – Belongs to RNA Polymerase family. Does not need a free 3' hydroxyl group, so it creates RNA primer strands at the initiation sites. DNA Ligase - Nicks occur in the developing molecule because the RNA primer is removed and synthesis proceeds in a discontinuous manner on the lagging strand. The final replication product does not have any nicks because DNA ligase forms a covalent phosphodiester linkage between 3'-hydroxyl and 5'-phosphate groups.

DNA Polymerase Each time a cell divides, DNA polymerase duplicates all of its DNA, and the cell passes one copy to each daughter cell. In this way, genetic information is passed from generation to generation. DNA polymerase is the most accurate enzyme. It creates an exact copy of your DNA each time, making less than one mistake in a billion bases. After it copies each base, it proofreads it and cuts it out if the base is wrong.

Lagging and leading strands

The Lagging Strand In DNA replication, the lagging strand is the DNA strand at the opposite side of the replication fork from the leading strand. It goes from 3' to 5' (these numbers indicate the position of the molecule in respect to the carbon atoms it contains). On the lagging strand, primase "reads" the DNA and adds RNA to it in short bursts. Pol III lengthens the bursts, forming Okazaki fragments. Pol I then "reads" the fragments, removes the RNA, and adds its own nucleotides (this is necessary because RNA and DNA use slightly different kinds of nucleotides). DNA ligase joins the fragments together.

Leading Strand Lagging Strand Okazaki Fragments (Discontinuous replication)

Lagging and leading strands

Telomeres and Telomerase The DNA of human telomeres comprises of an extremely short and simple sequence of nucleotides, TTAGGG,[28,29] repeated over and over. An important characteristic to note is that the telomere strand is particularly rich in Guanine(G) bases.

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