Bio 55 Ch. 10-11 MA post PDF

Microbial Genetics Big Picture: Genetics Alteration of bacterial genes and/or gene expression  Cause of disease  Prevent disease treatment  Manipulated for human benefit © 2012 Pearson Education, Inc.  Chapter 10-11: Microbial Genetics The Structure and Replicat...

Microbial Genetics Big Picture: Genetics Alteration of bacterial genes and/or gene expression  Cause of disease  Prevent disease treatment  Manipulated for human benefit © 2012 Pearson Education, Inc.  Chapter 10-11: Microbial Genetics The Structure and Replication of Genomes – Genetics – study of inheritance and inheritable traits as expressed in an organism’s genetic material – Genome – the entire genetic complement of an organism – Includes ALL its genes and nucleotide sequences Copyright © 2009 Pearson Education Inc., publishing as Pearson Benjamin Cummings Nuclear (eukaryotic) chromosomes – Typically have more than one chromosome per cell – are linear --within membrane-bound nucleus – Eukaryotic cells often have two copies of each chromosome (diploid) – Histones associated for packaging chromosome Copyright © 2009 Pearson Education Inc., publishing as Pearson Benjamin Cummings Extranuclear DNA of eukaryotes DNA molecules of mitochondria and chloroplasts ---are circular & resemble chromosomes of prokaryotes – Only codes for about 5% of RNA and proteins – Nuclear DNA codes for 95% of RNA and proteins –strong evidence for endosymbiosis theory A few fungi and protozoa carry plasmids Copyright © 2009 Pearson Education Inc., publishing as Pearson Benjamin Cummings Prokaryotic chromosomes – DNA, associated proteins (archaea have histones like eukaryotes) & RNA, packaged in 1-2 distinct chromosomes – single copy of each chromosome (haploid) – Typically circular chromosome – in nucleoid Copyright © 2009 Pearson Education Inc., publishing as Pearson Benjamin Cummings  Plasmids – Small, typically circular, molecules of DNA that replicate independently – Carry information required for their own replication, and often for one or more cellular traits – Not essential for normal metabolism, growth, or reproduction – Can confer survival advantages – Many types: – Fertility factors – Resistance factors – Bacteriocin factors – Virulence plasmids Copyright © 2009 Pearson Education Inc., publishing as Pearson Benjamin Cummings  The Structure and Replication of Genomes Helpful comparison chart [INSERT TABLE 7.1] Copyright © 2009 Pearson Education Inc., publishing as Pearson Benjamin Cummings DNA Replication – An anabolic polymerization --requires monomers and energy – Triphosphate deoxyribonucleotides serve both functions – Key to replication is complementary structure of the two strands – Replication = semiconservative – new strands composed of one original strand and one daughter strand Copyright © 2009 Pearson Education Inc., publishing as Pearson Benjamin Cummings  The Structure and Replication of Genomes (a) Each deoxyribonucleotide is made up of a sugar called deoxyribose, a phosphate group, and a nitrogenous base—in this case, guanine. (b) The five carbons within deoxyribose are designated as 1ʹ, 2ʹ, 3ʹ, 4ʹ, and 5ʹ. [INSERT FIGURE 7.4] Copyright © 2009 Pearson Education Inc., publishing as Pearson Benjamin Cummings  Figure 10.12- 13 DNA polymerization - Phosphodiester bonds form between the phosphate group at 5ʹ carbon of one nucleotide and the OH DNA nitrogenous bases  two-ringed of the 3ʹ carbon in the next nucleotide. DNA purines (A & G) & single-ringed pyrimidines is made in the 5’  3’ direction (C & T). Thymine is unique to DNA.  Figure 10.16 Watson and Crick proposed the double helix model for DNA. (a) The sugar-phosphate backbones are on the outside of the double (b) The two DNA strands are antiparallel to each other. (c) The direction of each strand is identified by numbering the carbons (1 through 5) in each sugar molecule. The 5ʹ end is the one where carbon #5 is not bound to another nucleotide; the 3ʹ end is the one where carbon #3 is not bound to another nucleotide. (d) Hydrogen bonds form between complementary nitrogenous bases on the interior of DNA. EXERCISE 49  DNA Replication – Initial processes in replication: DNA polymerase binds to each strand and adds nucleotides to hydroxyl group at 3′ end of nucleic acid DNA polymerase III Replicates DNA only 5′  3′ (3’ is growing end) – Because strands are antiparallel, – Leading strand synthesized continuously – Lagging strand synthesized discontinuously Copyright © 2009 Pearson Education Inc., publishing as Pearson Benjamin Cummings  The Structure and Replication of Genomes Copyright © 2009 Pearson Education Inc., publishing as Pearson Benjamin Cummings  DNA Replication REVIEW Two replication forks are formed by the opening of the double-stranded bidirectional circular DNA at the origin Two replication forks are formed by the opening of the double-stranded DNA at the origin Copyright © 2009 Pearson Education Inc., publishing as Pearson Benjamin Cummings  DNA Replication: REVIEW 1. Topoisomerase relaxes the supercoiled chromosome at the origin of replication, 2. Two replication forks formed 3. Helicase separates the DNA strands (breaks hydrogen bonds) 4. DNA is coated by single-stranded binding proteins (SSBPs) to keep the strands separated. 5. RNA primase lays RNA primer complementary to the parental strand 6. DNA polymerase III- elongates primer by adding nucleotides to the 3ʹ-OH end (in both directions). a. leading strand- DNA is synthesized continuously (5’  3’) b. lagging strand- DNA is synthesized in Okazaki fragments. 7. DNA Polymerase I - has exonuclease activity; removes RNA primers within the lagging strand 8. Ligase joins Okazaki fragments Copyright © 2009 Pearson Education Inc., publishing as Pearson Benjamin Cummings Important Enzymes in DNA Replication, Expression, and Repair REVIEW Copyright © 2009 Pearson Education Inc., publishing as Pearson Benjamin Cummings Growth of Microbial Populations When do bacteria replicate? Replication occurs prior to cell division (fission) replicate their DNA continuously as they grow and prepare to divide Copyright © 2009 Pearson Education Inc., publishing as Pearson Benjamin Cummings Which of these steps is LAST in the order of events in DNA replication? a. Primase lays down the RNA primer. b. DNA polymerase III synthesizes the new strand. c. DNA helicase unwinds the double helix. d. DNA ligase seals the nick. © 2012 Pearson Education, Inc.  The Replication of Eukaryotic DNA –Similar to bacterial replication –Some differences Only occurs during S-phase of cell cycle Use’s four DNA polymerases Thousands of replication origins Shorter Okazaki fragments Plant and animal cells methylate only cytosine bases Copyright © 2009 Pearson Education Inc., publishing as Pearson Benjamin Cummings  Figure 11.2 Flow of genetic information  Central Dogma The central dogma states that DNA encodes messenger RNA, which, in turn, encodes protein. Phenotype is determined by the specific genes expressed under specific conditions. Cells may have the same genotype, but they may exhibit a wide range of phenotypes resulting from differences in patterns of gene expression in response to different environmental conditions. Gene Function The Relationship Between Genotype and Phenotype – Genotype – set of genes in the genome – Phenotype – physical features and functional traits of the organism The Transfer of Genetic Information – Transcription – information in DNA is copied as RNA sequences – Translation – polypeptides synthesized from RNA sequences – Central dogma of genetics – DNA transcribed to RNA – RNA translated to form polypeptides Copyright © 2009 Pearson Education Inc., publishing as Pearson Benjamin Cummings  Figure 10.24 Genotype vs. Phenotype Gene expression changes as a response to environmental factors:  Nutritional, physical Streptococcus mutans (oral bacterium)- causes dental plaque; produces a sticky slime layer that adheres to teeth. Slime layer genes expressed in presence of sucrose (table sugar). Serratia marcescens (Gram-; associated with hospital-acquired infections [HAI]) have the gene for red pigment. However, this gene is expressed at 28 °C (left) but not at 37 °C (right). (credit: modification of work by Ann Auman) This Photo by Unknown Author is licensed under CC BY Gene Function [INSERT FIGURE 7.7] Copyright © 2009 Pearson Education Inc., publishing as Pearson Benjamin Cummings The Events in Transcription – Four types of RNA transcribed from DNA – RNA primers – mRNA – rRNA – tRNA – Occur in nucleoid of prokaryotes – Three steps – Initiation – Elongation – Termination Copyright © 2009 Pearson Education Inc., publishing as Pearson Benjamin Cummings Figure 7.9a The events in the transcription of RNA in prokaryotes. 1a RNA polymerase attaches nonspecifically to DNA and RNA polymerase travels down its length until 5′ 3′ DNA it recognizes a promoter 3′ 5′ sequence. Sigma factor enhances promoter Promoter Sigma factor Terminator recognition in bacteria. Attachment of RNA polymerase 1b Upon recognition of the “Bubble” promoter, RNA polymerase 3′ unzips the DNA molecule 5′ beginning at the promoter. 3′ 5′ Template Unzipping of DNA, movement of RNA polymerase DNA strand Initiation of transcription Copyright © 2009 Pearson Education Inc., publishing as Pearson Benjamin Cummings Figure 7.9b The events in the transcription of RNA in prokaryotes. “Bubble” 2 Triphosphate ribonucleotides align with their DNA 3′ 5′ 3′ complements and RNA 3′ 5′ polymerase links them together, synthesizing RNA. Growing RNA molecule No primer is needed. The (transcript) 5′ triphosphate ribonucleotides 5′ 3′ also provide the energy required for RNA synthesis. G Template C 5′ DNA 3′ strand Elongation of the RNA transcript Copyright © 2009 Pearson Education Inc., publishing as Pearson Benjamin Cummings Figure 7.9c The events in the transcription of RNA in prokaryotes. 3a 3b C Copyright © 2009 Pearson Education Inc., publishing as Pearson Benjamin Cummings Transcriptional differences in eukaryotes – RNA transcription occurs in the nucleus – Transcription also occurs in mitochondria and chloroplasts – Three types of RNA polymerases – Numerous transcription factors – mRNA processed before translation – Capping – Polyadenylation – Splicing Copyright © 2009 Pearson Education Inc., publishing as Pearson Benjamin Cummings Translation – Process whereby ribosomes use the genetic information of nucleotide sequences to synthesize polypeptides – Participants in translation: Ribosomal RNA (rRNA): integral part of ribosomes Transfer RNA (tRNA): transports amino acids during protein synthesis Messenger RNA (mRNA): carries coded information from DNA to ribosomes Copyright © 2009 Pearson Education Inc., publishing as Pearson Benjamin Cummings Functions of RNA in Protein Synthesis Copyright © 2009 Pearson Education Inc., publishing as Pearson Benjamin Cummings Figure 7.13 Prokaryotic mRNA molecules typically lack introns and exons. Copyright © 2009 Pearson Education Inc., publishing as Pearson Benjamin Cummings Figure 7.15 Ribosomal structures. Copyright © 2009 Pearson Education Inc., publishing as Pearson Benjamin Cummings The genetic code [INSERT FIGURE 7.11] Copyright © 2009 Pearson Education Inc., publishing as Pearson Benjamin Cummings Transfer RNA [INSERT FIGURE 7.13] Copyright © 2009 Pearson Education Inc., publishing as Pearson Benjamin Cummings The ribosome [INSERT FIGURE 7.15] Copyright © 2009 Pearson Education Inc., publishing as Pearson Benjamin Cummings Translation – Three stages – Initiation - initiator tRNA carrying N-formyl-methionine – Elongation – Termination – All stages require additional protein factors – Bacteria uses tRNA carrying N-formyl-methionine as initiator – Initiation and elongation require energy (GTP) Copyright © 2009 Pearson Education Inc., publishing as Pearson Benjamin Cummings Translation- elongation [INSERT FIGURE 7.17] Copyright © 2009 Pearson Education Inc., publishing as Pearson Benjamin Cummings Polyribosomes in prokaryote [INSERT FIGURE 7.18] In prokaryotes, multiple RNA polymerases can transcribe a single bacterial gene while numerous ribosomes concurrently translate the mRNA transcripts into polypeptides. In this way, a specific protein can rapidly reach a high concentration in the bacterial cell. Copyright © 2009 Pearson Education Inc., publishing as Pearson Benjamin Cummings Termination of translation –Release factors somehow recognize stop codons and modify ribosome to activate ribozymes, which sever polypeptide from final tRNA –Ribosome dissociates into subunits –Polypeptides released at termination may function alone or together- must fold Copyright © 2009 Pearson Education Inc., publishing as Pearson Benjamin Cummings Regulation of Genetic Expression – 75% of genes are expressed at all times – Other genes are regulated so they are only transcribed and translated when cells need them –Allows cell to conserve energy – Regulation of protein synthesis –Typically halts transcription –Can stop translation directly Copyright © 2009 Pearson Education Inc., publishing as Pearson Benjamin Cummings Gene Function: regulation of genetic expression –An operon consists of a promoter and a series of genes –Some operons are controlled by a regulatory element called an operator Copyright © 2009 Pearson Education Inc., publishing as Pearson Benjamin Cummings operons – Inducible operons must be activated by inducers – Lactose operon – Regulates lactose catabolism – Repressible operons are transcribed continually until deactivated by repressors – Tryptophan operon – Regulates tryptophan synthesis Animation: Operons Overview Copyright © 2009 Pearson Education Inc., publishing as Pearson Benjamin Cummings The lac operon; inducible [INSERT FIGURE 7.20] Animation: Operons Induction Copyright © 2009 Pearson Education Inc., publishing as Pearson Benjamin Cummings Figure 7.21 CAP-cAMP enhances lac transcription. Copyright © 2009 Pearson Education Inc., publishing as Pearson Benjamin Cummings Figure 7.22 The lac operon, an example of an inducible operon. Copyright © 2009 Pearson Education Inc., publishing as Pearson Benjamin Cummings Trp operon: repressible [INSERT FIGURE 7.21] Animation: Operons Repression Copyright © 2009 Pearson Education Inc., publishing as Pearson Benjamin Cummings Gene Function [INSERT TABLE 7.3] Copyright © 2009 Pearson Education Inc., publishing as Pearson Benjamin Cummings Gene Function Regulation of Genetic Expression – RNA molecules can control translation – Regulatory RNAs can regulate translation of polypeptides – microRNAs (miRNAs) – Bind complementary mRNA and inhibit its translation – Small interfering RNA (siRNA) – RNA molecule complementary to a portion of mRNA, tRNA, or DNA that binds and renders the target inactive – Riboswitch – RNA molecule that changes shape to help regulate translation Copyright © 2009 Pearson Education Inc., publishing as Pearson Benjamin Cummings Mutations of Genes Mutation – change in the nucleotide base sequence of a genome Rare event Almost always deleterious Rarely leads to a protein having a novel property that improves ability of organism and its descendents to survive and reproduce Mutagens increase rate of mutations 10-100 times Copyright © 2009 Pearson Education Inc., publishing as Pearson Benjamin Cummings Mutations of Genes Types – Point mutations (most common) – one base pair is affected insertions, deletions, and substitutions (silence, missense, nonsense) – Frameshift mutations – nucleotide triplets after the mutation are displaced – Insertions and deletions – Gross mutations – Inversions, duplications, transpositions Copyright © 2009 Pearson Education Inc., publishing as Pearson Benjamin Cummings Figure 7.24 The effects of the various types of point mutations. Copyright © 2009 Pearson Education Inc., publishing as Pearson Benjamin Cummings  Mutagens – Radiation –Ionizing radiation – induces breaks in chromosomes –Nonionizing radiation – induces pyrimidine dimers – Chemical Mutagens –Nucleotide analogs – disrupt DNA and RNA replication and cause point mutations –Nucleotide-altering chemicals – result in base-pair substitution mutations and missense mutations –Frameshift mutagens – result in nonsense mutations Copyright © 2009 Pearson Education Inc., publishing as Pearson Benjamin Cummings Mutations: effect of nucleoside analog [INSERT FIGURE 7.24] Copyright © 2009 Pearson Education Inc., publishing as Pearson Benjamin Cummings Mutations of Genes: Insertion of nucleotides Polymerase makes mistake where chemical is present [INSERT FIGURE 7.25] Copyright © 2009 Pearson Education Inc., publishing as Pearson Benjamin Cummings Identifying Mutants, Mutagens, and Carcinogens – Mutants – descendents of a cell that does not successfully repair a mutation – Wild types – cells normally found in nature – Methods to recognize mutants – Positive selection – Negative (indirect) selection – Ames test Copyright © 2009 Pearson Education Inc., publishing as Pearson Benjamin Cummings Positive selection: Copyright © 2009 Pearson Education Inc., publishing as Pearson Benjamin Cummings Replica plating: negative selection Copyright © 2009 Pearson Education Inc., publishing as Pearson Benjamin Cummings Ames test Purpose: [INSERT FIGURE 7.29] How to: Copyright © 2009 Pearson Education Inc., publishing as Pearson Benjamin Cummings  Genetic Recombination and Transfer Exchange of nucleotide sequences often mediated by DNA segments composed of homologous sequences Recombinants – cells with DNA molecules that contain new nucleotide sequences Vertical gene transfer – organisms replicate their genomes and provide copies to descendants Copyright © 2009 Pearson Education Inc., publishing as Pearson Benjamin Cummings  Horizontal Gene Transfer Among Prokaryotes – Horizontal gene transfer – donor cell contributes part of genome to recipient cell – Three types –Transformation –Transduction –Bacterial conjugation Copyright © 2009 Pearson Education Inc., publishing as Pearson Benjamin Cummings  Transformation: The Griffith Experiment – Transforming agent = DNA; -one of conclusive pieces of proof that DNA is genetic material – Competent cells; results from alterations in cell wall & cytoplasmic membrane that allow DNA to enter cell Copyright © 2009 Pearson Education Inc., publishing as Pearson Benjamin Cummings  Transduction Generalized – transducing phage carries random DNA segment from donor to recipient Relies on lytic cycle Animation Generalized Transduction Specialized – only certain donor sequences are transferred Relies on lysogenic cycle Animation: Specialized Transduction Copyright © 2009 Pearson Education Inc., publishing as Pearson Benjamin Cummings  Conjugation Animation: Conjugation F Factor Animation: Conjugation Overview Copyright © 2009 Pearson Education Inc., publishing as Pearson Benjamin Cummings HFR cells Animation: HFR Conjugation [INSERT FIGURE 7.34] Copyright © 2009 Pearson Education Inc., publishing as Pearson Benjamin Cummings When is the lac operon turned on/transcribed? A. when lactose is absent B. When lactose is present C. When the cell needs an energy source D. When glucose is absent Copyright © 2009 Pearson Education Inc., publishing as Pearson Benjamin Cummings