Lecture16S10 - BIS101/Engebrecht Lecture16 5/6/10 We...

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BIS101/Engebrecht Lecture16 5/6/10 We reviewed how transcription is initiated and elongates. How does it terminate? Termination: After several rounds of adding nucleotides, sequences residing in the RNA molecule itself trigger transcriptional termination, which is the dissociation of the RNA polymerase from the DNA and the release of the RNA chain . Two mechanisms exist for transcriptional termination (please see Handout15 ). The first is rho-independent (sometimes referred to as intrinsic terminators) termination. This requires the presence of 2 signs within the RNA molecule. First, there is a short inverted repeat (~20 nt) that will form a hairpin loop, this is followed by a stretch of As and Us. As soon as this region of the RNA is transcribed, the hairpin forms, and in combination with the weak AU pairing immediately adjacent to the hairpin, pulls the polymerase from the DNA molecule. Rho-dependent termination uses the protein Rho to help mediate dissociation of the RNA polymerase from the DNA template. The Rho protein binds a site on the RNA, called the rut site. Upon binding rho unwinds RNA from DNA template, causing the RNA polymerase to dissociate. Approximately 50% of the genes use rho- independent termination and 50% use rho-dependent termination. Gene density: E. coli has a very compact genome. In fact, many genes are transcribed from opposite strands of the dsDNA and can even overlap. This is why transcriptional termination signals are important. Please consider the phenotype of rho mutants. Summary: Transcription = RNA from DNA. Please understand the directionality of transcription, that specific signals or road signs are used to tell the RNA polymerase where to start transcription and where to end transcription . In E. coli, one RNA polymerase makes mRNA, tRNA, rRNA and snRNAs (next week we will see that eukaryotes using multiple RNA polymerases). The Genetic Code: The code was deciphered by a combination of genetics and biochemistry. What people wanted to know was how the nucleotide sequence dictated the protein sequence. Work done by Yanofsky provided strong evidence for the fact that the code was 1) nonoverlapping (since a single mutation only affected a single amino acid) and 2) that there was colinearity between the nucleotide sequence and the amino acid sequence. He accomplished this be isolating a large number of mutations in the trpA gene, encoding tryptophan synthetase, mapping them using recombination and then sequencing the resulting proteins. For example, if the order of the mutants was: Gene map _____1 _____2 ____3 ______4 ___ WT Tyr Leu Thr Gly Mutant 1 Lys Leu Thr Gly Mutant 2 Tyr Arg Thr Gly Mutant 3 Tyr Leu Ile Gly Mutant 4 Tyr Leu Thr Arg He found that the change in amino acid correlated with the position of the change in the protein. The triplet code hypothesis was formulated based on simple mathematics about the
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This note was uploaded on 07/02/2010 for the course BIS 101 taught by Professor Simonchan during the Spring '08 term at UC Davis.

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Lecture16S10 - BIS101/Engebrecht Lecture16 5/6/10 We...

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