Baker lecture Notes

Baker lecture Notes - Bisc 320, Molecular Biology lecture...

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Bisc 320, Molecular Biology lecture notes, Fall, 2007. Week 8: Molecular Genetics Some notes on: Conservative site-specific recombination and transposons See also, Watson, et al., chapter 11. > 40% of severe hemophilia A is caused by an inversion in exon #22 in the 26 exon factor VIII gene. Leukemias: reciprocal exchange of exons between chromosomes. Transposon terminal d.s. DNA sequences: IR: Inverted repeats (palindromes) allow inversions to occur via: IR--------IR DR: Direct repeats: deletions (excisions) and insertions occur via DR---------DR Figs 11-1, 11-11-2, 11-3 DNA-protein synaptic complexes form at pairs of recombinant recognition sequences. Conservative site-specific recombinases: Table 11-1, Fig 11-5 1. Serine type : cause d.s. breaks ; intermediate is a non-Holliday , non-heteroduplex, structure having a free 3’OH on each d.s. DNA piece, with a monomeric serine protein hooked to the 5’ end of the two DNA strands. No ATP needed. Staggered breaks (nicks) in each of the d.s. DNAs. Four recombinase monomers are used in recombination between two DNA duplexes. Recombination is via swapping of left and right s.s. overhangs of the four ends. Fig 11-6 a . Salmonella Hin invertase can invert a segment of bacterial genomic DNA. Before flip, in parenthesis: (hixL hin P hixR) flijB flijA, is in the “on” rightward-xcription configuration, that allows flijA expression. If there is a flip to the “off” configeration, transcription from P is then to the left. hixL and hixR are 60 BP each and palindromic to each other. Fis (f actor for i nversion s timulation) bends DNA. 4 Hin/2 Fis complex with HU protein forms at hix sites. (HU, unlike IHF is sequence independent.) flijB encodes H2 flagellin; flijA encodes repressor of distant flagellin gene. “On” allows only H2-type of flagella; “off” causes H1 flagella to appear on bacterial surfaces. This switch may elude the mammalian immune system by changing the type of flagella exposed on bacterial surfaces. Food poisoning. Fig 11-11, 11-12, 11-13 2. Tyrosine type : cause s.s breaks ; intermediate is a Holliday structure with a protein monomer hooked via tyrosine to the 3’ ends of nicks. Rotation of structure produces recombinant. Four monomers are used in recombination between two DNA duplexes. An acidic DDE (aspartate, aspartate, glutamate) motif is often found in tyrosine recombinase proteins, such as MuA, TN5, and RSV integrase: NH2---DNA binding site---DDE---protein-protein binding site—COOH Fig 11-7 a . P1 bacteriophage Cre protein acts at pairs of lox sites, creating a deletion of the lox sequences and the intervening DNA: Four subunits of Cre create a Holliday structure, prior to the deletion event. Fig 11-8 b . Lambda bacteriophage: lysogenic state vs lytic growth after viral infection. After infection, linear d.s. lambda DNA circularizes via 12/12 s.s. DNA overlap. Integration of the new lambda DNA circle is via a Holliday structure, with crossing over at: PP’ X BB’
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There is sequence homology between 7 out of the 15 base pairs at this crossover site. The E. coli lysogenic genome then is:
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This note was uploaded on 02/28/2008 for the course BISC 320L taught by Professor Baker,aparicio during the Fall '07 term at USC.

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Baker lecture Notes - Bisc 320, Molecular Biology lecture...

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