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Genetics Study Notes.docx

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Western University
Biology 2581B
Jim Karagiannis

Genetics Study Notes DNA, the Molecule of Heredity and Biological Information INFORMATION: is “that which reduces uncertainty” How do you quantify information? Uncertainty = log (2), where M is the # of possible symbols. Maximum information content of any sequence = L[log (M2], where L is the length of the sequence - For DNA: 4 possible symbols A,G,C, or T - Uncertainty= log (4) =2 bits 2 - Bits is the unit for information How DNA was experimentally shown to carry information Goal: figure out what in the cellular debris was actually conveying the message (information) - First they showed that this whole system was independent of the mouse. Could do the same thing in vitro. - Started purifying cellular debris into different components – DNA, protein and, lipids - Found it was the DNA component that could transform cells to S form. - Some scientists were skeptics, and said it wasn’t purified. And therefore they couldn’t know for sure. - Used enzymes to destroy certain components. DNA is the Genetic Material What was the phage injecting? DNA or protein? - Two experiments - 1. Take phage, infect E.coli, grow them in media that has radioactive P, - 2. Same thing, but phages labeled with radioactive S. - Why P-32 and S-35? - P is found in DNA, not protein, Sulfur is found in protein but NOT DNA. - 1. Infect E.coli- create new phage. DNA is labeled. Radioactivity was not associated with the ghosts. It was found inside the cells. Phage are introducing DNA into the cell… - 2. Same routine- this time radioactivity is associated with ghosts and not bacteria. The Eukaryotic Chromosome  Grooves (major and minor) are as a result of the geometry of the base pairings. o The angle between the glycosidic bonds of the minor groove is 120 degrees. The angle between the glycosidic bonds of the major groove is 240 degrees.  B-DNA represents only one possible conformation that a DNA double helix can form.  Other forms of DNA also exist e.g. A-DNA. However, they do not form under physiological conditions and are thus not biologically relevant. Only B and Z are physiologically relevant.  Biological Function of Z-DNA  The Biological role of Z-DNA remains mysterious, but some evidence suggests that Z-DNA may have a functional role within cells: o Z-DNA is formed transiently in association with transcription o Several proteins identified with highly specific Z-DNA binding activities e.g. Vaccinia virus E3L protein: essential for virulence o Antibodies to Z-DNA bind transcriptionally active regions Flexibility in helical structure  A crucial property of the double helix is its ability to separate the two strands without disrupting covalent bonds. This makes it possible for the strands to separate and reform under physiological conditions. o This is important for the processes of DNA replication, transcription, and also for DNA repair mechanisms.  Base Flipping o Enzymes involved DNA repair may scan for DNA lesions by flipping out bases Flexibility in DNA Organization  Linear, circular, single stranded or double stranded. Lots of variety. Flexibility in DNA Topology  Topology: 3d organization of a molecule in space.  Topologically constrained: if you were to denature the strands, you wouldn’t be able to separate strands- they are interwoven  No Topological Constraint: If you denature the strands you could separate strands  For a constrained double helix, torsional stress introduces… supercoiling  DNA does (-) supercoiling, makes it more compact  Supercoiling can be induced if the DNA molecule is underwound before the circle is made.  This destabilizes the helix (260 bp/23 turns= 11.3 bp/turn).  There are now two ways to stabilize the DNA molecule: 1) partially separate the strands, or 2) introduce supercoils. Why do Living Cells store DNA with Negative Superhelicity?  Many advantages in storing energy in this way  Can be called on when needed for processes that require strand separation e.g. DNA replication, transcription DNA Packaging in Eukaryotes • Average DNA in human chromosome = 3cm long • 46 chromos = almost 2 m of DNA per cell • Yet fits in nucleus that is 10 microns in diameter! • Clearly DNA is very efficiently packed into the chromosome • HOW?? • What is Chromatin? • The complex of DNA, chromosomal proteins, and other chromosomal constituents isolated from nuclei • Chemical analysis of chromatin reveals: • Primarily DNA and protein with some RNA • Proteins fall into two classes: • Histones • Non-histone proteins • Histones – small proteins with basic, positively charged amino acids lysine and arginine • Bind to and neutralize negatively charged DNA • Make up half of all chromatin protein by weight • Five types: H1, H2A, H2B, H3, and H4 • Core histones make up nucleosome: H2A, H2B, H3, H4 • DNA and histone synthesis regulated so that both are synthesized together during S phase • High level of similarity of histones among diverse organisms • Nonhistone proteins are a heterogeneous group: • Half of proteins in chromatin are nonhistone • Large variety of functions • Scaffold – backbone of chromosome • DNA replication – e.g., DNA polymerases • Chromosome segregation – e.g., motor proteins of kinetochores • Transcriptional regulation • Unknot and disentangle DNA molecules-topoisomerases The nucleosome: the fundamental unit of chromosomal packaging arises from the association of DNA with histones • Chromatin fibers with beads having diameter of about 10 nm and strings having diameter of 2 nm • Bead is a nucleosome with about 160 bp of DNA wrapped twice around a core of 8 histones. • 40 bp of DNA link individual nucleosomes together • The wrapping of DNA around the histone core stores negative superhelicity • SINCE negatively supercoiled DNA favors DNA unwinding, the removal of nucleosomes will: • Increase access to DNA • Promote DNA unwinding of nearby DNA sequences • IMPORTANT FOR: DNA replication, transcription • Higher order Chromosome Structure • With H1 the coils coil around each other- to make 30 nm (3 nucleosomes across) fibers • Radial loop scaffold model for higher levels of compaction • Each loop contains 60-100 kb of DNA tethered by nonhistone scaffold proteins Unineme Model For Chromosome Structure  There is just one DNA double helix extending from one chromosome end to the other  To support this they carefully unwound a chromosome and found it to be one giant linear model  Second experiment  measured how long it took to recoil. This is proportional to length Gene and Genome Structure DNA: Information Storage and Retrieval DNA stores the biological information to create a diverse range of • Proteins  cell types  tissues  organisms • Advantages: • Ease of storage (large quantity of data) • Can be copied reliably • DNA stores information “digitally” • A C T G • Analogous to storing electronic data such as music Central Dogma of Molecular Biology and Genetics: DNA  RNA  Protein What is a gene? • The basic unit of biological information • A specific segment of DNA at a specific location in the genome (on a region of a chromosome) that serves as a unit of function • Encodes RNA or protein Anatomy of a eukaryotic gene Anatomy of a eukaryotic gene 3’ 5’ 3’ Coding strand: • similar 5’  3’ sequence as RNA • Sense , non-template, or Crick strand Non-coding strand: • used as a template to transcribe RNA • Antisense, template, or Watson stran
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