MICROBIO 310 1st Edition Lecture 8 Outline of Last Lecture I. 4.6 Electron Donors and Electron AcceptorsII. 4.7 Energy-Rich Compounds and Energy StorageIII. 4.8 Glycolysis IV. 4.9 Respiration and Electron Carriers V. 4.10 The Proton Motive Force VI. 4.11 The Citric Acid Cycle VII. 4.12 Catabolic Diversity VIII. 4.14 Biosynthesis of Amino Acids and NucleotidesIX. 4.15 Biosynthesis of Fatty Acids and LipidsOutline of Current Lecture I. 6.1 Macromolecules and GenesII. 6.2 The Double HelixIII. 6.3 SupercoilingThese notes represent a detailed interpretation of the professor’s lecture. GradeBuddy is best used as a supplement to your own notes, not as a substitute.IV. 6.4 Chromosomes and Other Genetic ElementsV. 6.5 The Escherichia coli ChromosomeVI. 6.6 Plasmids: General PrinciplesVII. 6.7 The Biology of PlasmidsVIII. 6.8 Templates and EnzymesIX. 6.9 The Replication ForkX. 6.10 Bidirectional Replication and the ReplisomeXI. 6.11 The Polymerase Chain ReactionXII. 6.12 Overview of TranscriptionXIII. 6.13 Sigma Factors and Consensus SequencesCurrent Lecture6.1 Macromolecules and Genes• DNA, RNA, Proteins: info macromolecules• Genetic information flow can be divided into three stages– Replication: DNA is duplicated – Transcription: information from DNA is transferred to RNA (first step in expression of a gene)• mRNA (messenger RNA): encodes polypeptides• tRNA (transfer RNA): plays role in protein synthesis• rRNA (ribosomal RNA): plays role in protein synthesis– Translation: information in RNA is used to build polypeptides– A gene is a unique recipe that codes for a protein but not all genes code for proteins- Central dogma of molecular biology – DNA to RNA to protein - Eukaryotes: each gene is transcribed individually- Prokaryotes: multiple genes may be transcribed together 6.2 The Double Helix• All cells and some viruses have DNA in double- stranded molecule• Two strands are antiparallel• Two strands have complementary base sequences (But they are not opposite to each other)– Adenine always pairs with Thymine – Guanine always pairs with Cytosine• Two strands form a double helix - Hydrogen bonds between DNA strands hold two strands together- Adenine–Thymine pair has two hydrogen bonds and Guanine–Cytosine pair has three hydrogen bonds– GC pairs are stronger than AT pairs- High heat breaks hydrogen bonds, causing denaturation (melting)- GC-rich DNA melts at higher temperatures than AT-rich DNA because it has more hydrogen bonds that need to be broken6.3 Supercoiling - Supercoiled DNA: DNA is further twisted to save space – Negative supercoiling: double helix is underwound – Positive supercoiling: double helix is overwound - Relaxed DNA: DNA has number of turns predicted by number of base pairs - Negative supercoiling is predominantly found in nature - DNA Gyrase: introduces supercoils into DNA6.4 Chromosomes and Other Genetic Elements- Genome: entire complement of genes in cell or virus- Chromosome: main genetic element in prokaryotes (complicated circle in prokaryotes)- Other genetic elements include virus genomes, plasmids, organellar genomes, and transposable elements- Viruses contain either RNA or DNA genomes – Can be linear or circular– Can be single or double stranded- Plasmids: replicate separately from chromosome– Great majority are double stranded– Most are circular (small circle)– Generally beneficial for the cell (e.g., antibiotic resistance)– NOT extracellular, unlike viruses (stay in the cytoplasm of cell)– Segment of DNA that can move from one site to another site on the same or a different DNA molecule - Chromosome is a genetic element with “housekeeping” genes – Presence of essential genes is necessary for a genetic element to be called a chromosome - Plasmid is a genetic element that is expendable and rarely contains genes for growth under all conditions (add ons-Don’t have essential genes)- Transposable Elements – Segment of DNA that can move from one site to another site on the same or a different DNA molecule (Jumping Genes)– Viewed as chromosomal parasites– Ex: Indian Corn– Inserted into other DNA molecules – Three main types: • Insertion sequences • Transposons• Special viruses 6.5 The Escherichia coli Chromosome- Escherichia coli is a useful model organism for the study of biochemistry, genetics, and bacterial physiology- The E. coli chromosome from strain MG1655 has been mapped using conjugation, transduction, molecular cloning, and sequencing• Some features of the E. coli chromosome– Many genes encoding enzymes of a single biochemical pathway are clustered into operons– Operons equally distributed on both strands– ~5 Mbp (megabase pairs) in size – ~40% of predicted proteins are of unknown function– Average protein contains ~300 amino acids– Insertion sequences (IS elements)6.6 Plasmids: General Principles - Plasmids: genetic elements that replicate independently of the host chromosome – Small circular or linear DNA molecules– Range in size from 1 kbp to >1 Mbp; typically less than 5% of the size of the chromosome – Carry a variety of nonessential, but often very helpful, genes – Abundance (copy number) is variable - A cell can contain more than one plasmid, but it cannot be closely related genetically due to plasmid incompatibility – Many incompatibility (Inc) groups recognized – Plasmids belonging to same Inc group exclude each other from replicating in the same cell but can coexist with plasmids from other groups • Some plasmids (episomes) can integrate into the cell chromosome; similar to situation seen with prophages• Removal (curing) plasmids from host cells can result from various treatments• Conjugative plasmids can be transferred between suitable organisms via cell-to-cell contact (pilus)– Conjugal transfer controlled by tra genes on plasmid6.7 The Biology of Plasmids• R plasmids– Resistance plasmids; confer resist– Many are conjugative (force transfer between cells)- In several pathogenic bacteria, virulence characteristics are encoded by plasmid genes - Virulence factors:– Enables pathogen to colonize– Enables pathogen to cause host damage – Hemolysin(break open red blood cells to get to iron)– Enterotoxin • Bacteriocins– Proteins produced by bacteria that inhibit or kill closely related species or even different strains (competitors) of the same species– Colicin, nisin• Genes encoding bacteriocins are often