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L9 4-15 Bacterial Genetics Bacteria’s genome contains: 1. One chromosome: circular with few associated proteins 2. Plasmids: smaller rings of circular DNA; not part of chromosome, so they are called accessory genes. - Self-replicating - Not necessary for the survival of the bacteria under normal conditions. - Can help bacteria if living in stressful conditions. Recombination of bacteria: [Note: In Eukaryotes = meiosis – fertilization – zygote = recombinant DNA] In Prokaryotes: Transformation: uptake of foreign DNA from surroundings - Bacteria have cell surface receptors that recognize and transport DNA from closely related species. - Result = recombinant; chromosome from two different cells. - Can alter bacteria’s genotype and phenotype. NOTE: THIS IS WHAT OUR NEXT LAB IS BASED ON…KNOW THIS! 1 L9 4-15 Transduction: transfer of DNA of one bacteria to another by bacteriophages (phages) as a result of phage’s life cycle Part of host DNA packaged into new “virus” capsid - phage injects its DNA into bacteria (host #1) - Host DNA is fragmented - Phage uses host DNA to make its own proteins and replicate its DNA - Host DNA is packaged into new viral capsid (not on purpose) - Lysis of host cell releasing many phages with host’s (#1) DNA - Phage with host’s #1 DNA infects bacterial host #2 - Recombination of host #1 and #2 DNA 2 L9 4-15 3 L9 4-15 Conjugation (bacterial “sex”): transfer of DNA through two bacteria temporarily joined via sex pilli (like a grappling hook) - “F” = “Fertility” - F+ cells = have F+ factor/gene carried on plasmid to form sex pilus and transfer DNA. - When “mating” of F+ cells and F- cells occurs, only F plasmid DNA is transferred and (not the chromosome) and the F- cell becomes an F+ cell. 4 L9 4-15 Control of Protein Production (in Eukaryotes) How much protein do we want synthesized? Don’t want to be wasteful! Modes of Regulation 1. Transcription - transcription factors (proteins) = “glue dot” - processing (cutting & pasting exons) - stability of mRNA in the cytoplasm 2. Translation - protein initiation factors = “glue dot” 5 L9 4-15 Let’s focus on the Transcriptional level of regulation: o Transcriptional control of mRNA production; happens on two levels: 1. (Pro. & Euk) Adjust the activity of enzymes already present by feedback inhibition. In general, feedback mechanisms operate by a(an) - Lack of substance turns on genes that code for enzymes to synthesize that substance. (positive feedback) - Abundance of substance will turn off genes that code for enzymes to build substance. (negative feedback) (see page 353 fig. 18.20) 2. (Pro. & Euk) Adjust the amount being made of certain enzymes by regulating expression of genes coding for enzymes Prokaryotes only - structural genes ("SG"): code for enzymes, structural proteins - regulatory genes: control the output of structural genes Regulatory examples – using Operon Models 6 L9 4-15 Operons a group of genes associated with other segments of DNA regulated as a whole by regulatory gene (not part of operon) The regulatory gene controls a part of the operon know as the operator gene = single “on/off” switch for the SG’s collectively Operons = operator + promoter + (few to many) SG The lac operon of E. coli: Regulates the hydrolysis of lactose (disaccharide) - catabolic Prokaryote example of INDUCIBLE OPERON = usually turned off by a repressor protein and needs to be induced (turned on) by specific molecule (lactose). E. coli in human’s digestive track; humans drink milk = E. coli goes to work = lac operon is induced - SG's for related enzymes adjacent on chromosome are activated == enzymes made - lactose metabolism uses these three enzymes • operator gene next to 3 SG's for lactose metabolism • promoter genes adjacent to operator • the operator and promoter do not code for mRNA Op This is the lac operon [ P + Op + SG1 + SG2 + SG3 ] In the presence of lactose: 7 L9 4-15 - lactose binds to the repressor protein - preventing repressor protein from binding to the operator - so transcription can continue unblocked This is an inducible operon: - it is induced to transcribe - by the presence of a molecule (lactose) - requiring enzymes for its metabolism o RNA Pol II attaches to promoter o synthesis of all three SG's starts a regulatory gene is located close by o this produces a repressor protein o which attaches to the operator o blocking RNA Pol II, preventing transcription 8 L9 4-15 The tryp operon of E. coli : Example of REPRESSIBLE OPERON = usually turned on until repressed by a specific molecule (ex. tryptophan) binds to regulatory protein. - if tryptophan is present, no need for synthesis o tryptophan (the co-repressor) binds to the repressor protein o which is then able to bind to the operator o blocking RNA Pol II, preventing transcription in absence of tryptophan o the repressor protein alone is unable to bind to operator o transcription is free to proceed this is a repressible operon - when a needed substance appears in environment - structural genes for its synthesis are no longer transcribed 9 L9 4-15 - saving the cell energy and materials So,… Lac operon and tryp operon both operate by negative feedback; stops expression of SG’s both are turned off by the active form of a repressor protein Lac operon also operates by positive feedback; induces expression of SG’s 10