MICRO-s10_15 - BIOL 240 General Microbiology Spring 2010 Rm...

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Unformatted text preview: 3/17/2010 BIOL 240: General Microbiology Spring 2010 Rm. 22-116 R, Mar. 18, 2010 22http://www.smccd.edu/accounts/staplesn/biol240/ 1. Pre-Lab Writeups: Be sure to prepare before each Monday’s labs (for Lab Writeups Be sure to prepare before each Monday labs (for BOTH BOTH Mon. & Wed.)!! (What? Why? How? are we doing in the lab??) (What? 2. Be sure to keep up with BY ARRANGEMENT HOURS!! They are REQUIRED for your grade!! -- average TWO documented hours/week. -- 3. MT #2 on Tuesday!!! See updated Study Guide!! • Review session today in Lab!! 12:45 pm!! 4. NEXT Tues.: water sample from OUTDOOR NEXT Tues water sample from OUTDOOR source!! source!! Get collection Kit in lab! (~50 ml) 5. QUIZ #4 Due TONIGHT!! 6. Notebook Checks “THIS week”, 3/16: Expt. 1-10, etc.!!! “THIS 1- 1. 1. Ch. 8: Describe the Central Dogma of molecular genetics and the three Central processes that drive the flow of genetic information in an organism. 2. Describe several properties of DNA and the process of DNA replication that contribute to properties DNA DNA’s central role as the hereditary material. 3. Draw a replication fork and label 5 enzymes involved in DNA replication. Describe the enzymes function of each enzyme. REVIEW: TODAY’s Objectives: Students should be able to... Students 1. Compare and contrast DNA Synthesis (Replication) with RNA Synthesis (Transcription). Why is it suitable that RNA is a less stable molecule than Why is it suitable that RNA is less stable molecule than DNA? (THINK: DNA? (THINK: Structure Function!!! ….. ALWAYS!!) [Make a table!] Function [Make table!] 2. ** List the three types of RNAs and their functions in protein synthesis. three How do they “read” the genetic code? 3. ** Compare the stages of Initiation, Elongation, and Termination in Initiation, Replication, Replication, Transcription and Translation. Indicate the direction of synthesis in each, and the names of the start and stop sites. [Make a table!] [Make table!] 1 3/17/2010 DNA replication is Semiconservative Bidirectional Semiconservative & Bidirectional • Replication results in two daughter DNA duplexes, •each with one each with one completely completely new strand, & •one old strand (parental strand) • = “SEMI-CONSERVATIVE” “SEMI- Figure 8.6 http://www.wehi.edu.au/education/wehihttp://www.wehi.edu.au/education/wehi-tv/dna/replication.html • Two replication forks move in opposite directions = •“Replication Bubble” 8.2) Transcription: RNA Synthesis RNA 1. DNA is transcribed to make RNA (AUGC) a) mRNA = messenger RNA translated translated to protein brings brings amino acid to ribos. b) tRNA = transfer RNA c) rRNA = ribosomal RNA ribosomal RNA makes up ribo; catalytic makes up ribo; catalytic 2. Transcription begins when RNA polymerase RNA binds to the PROMOTER sequence PROMOTER 3. Transcription proceeds in the 5′ 3′ direction (same in ALL nucleic acid synthesis!) • new nucleotides added to the 3’ hydroxyl group on ne added to the 3’ gro on ribose ribose in the growing strand 4. Transcription stops when it reaches the Terminator Sequence (often many U’s or “hairpin loop”) • New RNA and Rpol fall off of DNA template. 2 3/17/2010 http://www.stolaf.edu/people/giannini/flash animat/molgenetics/transcription.swf http://vcell.ndsu.nodak.edu/animations/ transcription/movie.htm Figure 8.7 RNA processing in Eukaryotes “Primary transcript” (coding exons + noncoding introns) Primary transcript (coding exons noncoding introns “Mature messenger RNA” (introns removed) (introns Exits nucleus to be translated by ribosomes Ribozymes!!! Ribozymes!!! Nuclear pores Figure 8.11 3 3/17/2010 8.3) Translation 1. mRNA is translated in Codons Codons – 3 nucleotide “words” nucleotide words 2. Translation of mRNA begins at the start start codon: AUG 3. Translation ends at a STOP (“ STOP (“nonsense”) codon codon: UAA, UAG, UGA – Do NOT encode an amino acid!!! Figure 8.2 The “Universal” Genetic Code Figure 8.8 • tRNA (transfer RNA) = adapter molecule that reads “3 letter” codons – via it’s complementary “Anticodon”! • Puts in the right amino acid in right order 3’ • Reads mRNA 5’ 3’, • synthesizes N C polypeptide • Amino Carboxy Carboxy! • Anticodon-Codon base Anticodonpairing http://www.lewport.wnyric.org/jwanamaker/ animations/Protein%20Synthesis.html 4 3/17/2010 A. Translation – Initiation Large 3’ 1. 5’ Small Figure 8.09.1 http://www.stolaf.edu/people/giannini/flashanimat/molgenetics/translation.swf http://highered.mcgrawhttp://highered.mcgraw-hill.com/sites/0072437316/student_view0/chapter15/animations.html# B. Translation - Elongation 2. Peptidyl-site eptidylAmino Acyl-site Acyl- 3’ 5’ Exit-site xit- E P A Figure 8.09.2 5 3/17/2010 Peptidyl Transferase 3. C N 3’ 5’ Translocation Figure 8.09.3 Newly Newly elongated peptide in A site, then then 4. Peptide bond!! C —N PP now in P site! 5’ 3 NT NT Translocation 3’ Figure 8.09.4 6 3/17/2010 5. Ribosome Translocates 3nt in the 3’ direction 3’ 5’ Translocation Translocation 3 NT Figure 8.09.5 Translation: Peptidyl Transfers & translocations continue 6. 3’ 5’ 3 NT….. Figure 8.09.6 Translocation 7 3/17/2010 C. Translation - Termination 7. Figure 8.09.7 5’ RF Last AA inserted BEFORE stop codon!! 3’ • No tRNA, but a protein protein “Release Factor Factor” enters empty A site at a stop codon Tsln - Termination/ Release 8. 5’ 3’ Figure 8.09.8 http://vcell.ndsu.nodak.edu/animations/translation/movie.htm 8 3/17/2010 ** Polyribosomes • Multiple ribos/ polypeptides translated per mRNA!!! per mRNA!!! • = POLYRIBOSOMES (in both Euk. & Prok.!) Also: • Tsln & Tscn simultaneous in prokaryotes!! (only!) • No nuclear membrane to separate! Figure 8.10 5’ 5’ mRNA Compare Repln, Tscn, Tsln: Process Replication Initiation Elongation Termination Terminator sequences, or end of chromosome (forks meet if circle) Tsc’l terminator (poly(poly-U, hairpin loop) Stop codon (nonsense codon): codon): UAA, UAG, UGA • Release Factor (= protein) At Origin: A/T- Dpol3, dNTPs, A/T- Dpol3 rich 5’ 3’ 3’ rich, NZs: Helicase, Primase, DPol3 RPol, NTPs, 5’ 3’ 3’ Transcription At Promoter: AT rich, NZ: RPol Translation At Start Codon Codon:(AUG), mRNA, AAAA-tRNA NZ: ribosome Ribosome, AAAA-tRNA’s (anticodons), N C (follows (follows mRNA 5’ 3’) 3’) 9 3/17/2010 8.4) 8.4) Regulation of Bacterial Gene Expression • Constitutive enzymes are expressed at a fixed rate (“always on”!) – “housekeeping” functions in cells • Other enzymes are expressed only as needed (“regulated enzymes”) enzymes – Repressible enzymes = turned off when not needed; usually ON. usually – Inducible enzymes = turned on only when needed; usually OFF. usually Repression • Prokaryotes: Related genes organized into OPERONS: OPERONS – Many “structural genes” Many “structural genes” (protein(protein-encoding) – Under the control of a SINGLE PROMOTER operator PROMOTER & operator (negative control sequence) – Polycistronic (polygenic) Polycistronic (polygenic) mRNA’s! mRNA’s! • Single mRNA transcript encodes encodes several proteins!! • NOT in EUKaryotes!! Figure 8.13 (2004) 10 3/17/2010 Lactose Catabolism; Trp Synthesis Operons: • OPERATOR = target/binding site on DNA • Bound by a REPRESSOR protein, to stop action of REPRESSOR RNA Polymerase ( genes OFF) genes • Negative Regulation! Figure 8.12. (2007) A. Lac Operon: Inducible Repressor protein usually bound to Operator (OFF!), unless lactose present. Figure 8.12.2 11 3/17/2010 Lac gene induction by Lactose SignalSignal-Bound Repressor = OFF (genes ON), in an INDUCIBLE system http://vcell.ndsu.nodak.edu/animations/lacOperon/movie.htm Figure 8.12.4 Positive Regulation of the lac operon lac Figure 8.15 1. Glucose is the preferred energy source in most cells. 2. For lactose and other alternative sugars to be consumed, glucose cannot be present. 3. With glucose absent, ATP ADP ADP AMP AMP cyclic cyclic AMP (starvation signal!!). 4. cAMP binds CAP (Catabolite Activator Protein), which helps RPol bind lac promoter CAP (Catabolite 5. Full LAC operon activation requires Lactose PRESENT, and Glucose ABSENT!!!. 12 3/17/2010 B. Trp Operon: Repressible Repressor protein NOT bound to Operator, unless excess TRP present. • TRP = “corepressor”; activates repressor • genes genes OFF when excess TRP Figure 8.13 Trp gene repression by excess TRP SignalSignal-Bound repressor = ON (genes OFF OFF), in a REPRESSIBLE system Figure 8.13 13 3/17/2010 Regulation of Gene Expression Faster growth on preferred C-source (glc) C- Figure 8.14 Delayed growth (Lag) on switching to secondary sugar, Lactose • Lact catabolic enzymes must be induced & synthesized!! Operon Regulation: Summary OPERON TYPE TYPE ACTIVITY of of Signal Signal Molecule Molecule Absent Absent Signal Molecule Molecule Present Present ― (lactose inducer) Catabolic, ~lac Repressor (Inducible) Protein Operon (transcription) (transcription) Anabolic, ~trp Repressor (Repressible) Protein Operon (transcription) (transcription) + OFF ― ON ON + (tryptophan corepressor) OFF 14 ...
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