Bacterial Genetics

The sum total of genetic material of a cell is referred to as the genome. The general location and forms of the genome

Chromosome Subdivided into basic informational packets called genes Procaryotic Histonelike proteins condense DNA Located in cytoplasm Usually circular Usually single chromosome

Chromosomes Eucaryotic Histone proteins condense DNA Located in nucleus Vary in number Can be diploid (sister chromatids) or haploid Appear elongate

Genes Part of DNA that encodes a protein Direct cell function Basic unit of heredity In general, linear sequence of nucleotides or codons with a fixed start and end point Genes--> chromosomes-->organisms

Genes encoded by DNA Three categories Genotype Phenotype Structural- encode proteins Regulatory- control gene expression Encode for RNA- rRNA or tRNA Genotype sum of all gene types Phenotype Collection or expression of traits encoded in an organism’s genotype

How genes effect heredity Genotype sum of all gene types Phenotype Collection or expression of traits encoded in an organism’s genotype

Central Dogma DNA RNA Protein replication translation transcription Reverse transcription

DNA Structure Nucleotide Double stranded helix Phosphate Deoxyribose sugar Nitrogenous base Double stranded helix Antiparallel arrangement

Nitrogenous bases Purines Adenine Guanine Pyrimidines Thymine Cytosine

Purines and pyrimidines pair (A-T or G-C) and the sugars (backbone) are linked by a phosphate. Three views of DNA structure

Replication Semiconservative Enzymes Leading strand Lagging strand Okazaki fragments

Semiconservative New strands are synthesized in 5’ to 3’ direction

Semiconservative replication of DNA synthesizes a new strand of DNA from a template strand. Simplified steps to show the semiconservative replication of DNA

Steps in replication Uncoiling- unwinding of DNA from histones Unzipping- breaking hydrogen bonds between base pairs allowing strands to separate Addition of nucleotides- each parent strand is used as the template for synthesis of daughter strands. Read 3’-5’, synthesis 5’-3’

Enzymes

Leading strand RNA primer initiates the 5’ to 3’ synthesis of DNA in continuous manner

Lagging strand Multiple Okazaki fragments are synthesized Okazaki fragments are ligated together to form one continuous strand Replication begins at the origin of replication

The steps associated with the DNA replication process. The bacterial replicon: a model for DNA Synthesis

Replication processes from other biological systems (plasmids, viruses) involve a rolling cycle. Simplified model of rolling circle DNA Replication

RNA Transcription Message RNA (mRNA)- protein Transfer RNA (tRNA) Ribosomal RNA (rRNA)

Transcription A single strand of RNA is transcribed from a template strand of DNA Template strand- transcribed Coding strand- nontranscribed RNA polymerase catalyzes the reaction In eucaryotes RNAP I- rRNA RNAP II- mRNA -->protein RNAP III-tRNA

Eucaryotic mRNA mRNA can have interruptions Exon expressed sequences Gene1 exon intron gene 1 exon Exon expressed sequences Intron intervening sequence Removed by splicesosomes. Splicesosome loops the intron into lariat shape ,excises them, and joins exons. Some introns become endonucleases.

The processing of pre-mRNA into mRNA involves the removal of introns.

RNA Thymidine is replaced by uracil Synthesis in 5’ to 3’ direction (with regard to RNA) The message contains a codon (three bases)

Transcription Initiates when RNAP recognizes a promoter region Two regions 1) 35 bp upstream from transcription start site 2) 10 bp upstream from transcription start site

Transcription Terminates when RNAP reaches terminators in sequence. These make RNAP stutter and fall off or pause and fall off

The synthesis of mRNA from DNA. The major events in mRNA synthesis

Transcription regulation Operons- coordinated set of genes which are regulated as a single unit. They are transcribed as a single unit. Types of operons Inducible Repressible

Regulator gene- gene that can repress the operon Operator- acts as on/off switch Structural gene- genes to be transcribed when the operon is on

Lactose operon (inducible) Breaks down lactose Normally in OFF position. Operon is controlled by the binding of the repressor to the operator Lactose binding to repressor causes conformation changes in the repressor Repressor dislodges from the operator RNAP binds and transcribes structural genes

When lactose levels fall: the repressor is free to bind the operator again In this system lactose is the inducer This operon is not inducible in the presence of glucose. Glucose is preferred over lactose as a carbon source

The regulation of sugar metabolism such as lactose involves repression in the absence of lactose, and induction when lactose is present. The lactose operon in bacteria

Repressible operon Operon is normally on Corepressor- normally the product of the operon. Turns operon off by binding and activating the repressor

The regulation of amino acids such as arginine involves repression when arginine accumulates, and no repression when arginine is being used. Repressible operon

mRNA Copy of a structural gene or genes of DNA Can encode for multiple proteins on one message Thymine is replaced by uracil The message contains codons Carries code for proteins

tRNA Copy of specific regions of DNA Cloverleaf structure Complimentary sequences form hairpin loops Amino acid attachment site Anticodon Participates in translation (protein synthesis) Delivers amino acid to growing polypeptide chain

Important structural characteristics for tRNA and mRNA. Characteristics of transfer and message RNA

rRNA Consist of two subunits (70S) A subunit is composed of rRNA and protein Participates in translation

Ribosomes bind to the mRNA, enabling tRNAs to bind, followed by protein synthesis. Summary of the flow of genetics

Codons Triplet code that specifies a given amino acid Multiple codes for one amino acid 20 amino acids Start codon Stop codons Redundant code allows for wobble

The codons from mRNA specify a given amino acid. The Genetic code

Representation of the codons and their corresponding amino acids.

Protein Translation Protein synthesis have the following participants mRNA tRNA with attached amino acid Ribosome

Participants involved in the translation process. The “players” in translation

Translation Ribosomes bind mRNA near the start codon (ex. AUG) tRNA anticodon with attached amino acid binds to the start codon Ribosomes move to the next codon, allowing a new tRNA to bind and add another amino acid Series of amino acids form peptide bonds Stop codon terminates translation

The process of translation. The events in protein synthesis

For procaryotes, translation can occur at multiple sites on the mRNA while the message is still being transcribed. Speeding up the protein assembly line in bacteria

Transcription and translation in eucaryotes Similar to procaryotes except AUG encodes for a different form of methionine mRNA code for one protein Transcription and translation are not simultaneous Pre-mRNA Introns Exons

Mutations Loss of bases Addition of bases Misincorporation of bases Wild-type: natural, nonmutated strain Mutant: harbors 1 or more mutations

Mutations Changes made to the DNA Spontaneous – random change Induced – chemical, radiation. Point – change a single base Nonsense – change a normal codon into a stop codon Back-mutation – mutation is reversed Frameshift – reading frame of the mRNA changes

Examples of chemical and radioactive mutagens, and their effects. Selected mutagenic agents and their effects

Ames test Salmonella culture (his-) cannot make histidine Minimal media (his-) + test reagent 6 colonies INDUCED + SPONTANEOUS MUTANTS Enriched media (his+) 18 colonies TOTAL BACTERIA Minimal media (his-) 3 colonies SPONTANEOUS MUTANTS

Effects of mutations Positive effects for the cell Allow cells to adapt Negative effects for the cell Loss of function Cells cannot survive

Recombination Sharing or recombining parts of their genome Conjugation Transformation Transduction

Conjugation Transfer of plasmid DNA from a F+ (F factor) cell to a F- cell An F+ bacterium possesses a pilus Pilus attaches to the recipient cell and creates pore for the transfer DNA High frequency recombination (Hfr) donors contain the F factor in the chromosome

Conjugation is the genetic transmission through direct contact between cells. Conjugation: genetic transmission through direct contact

Transformation Nonspecific acceptance of free DNA by the cell (ex. DNA fragments, plasmids) DNA can be inserted into the chromosome Competent cells readily accept DNA

DNA released from a killed cell can be accepted by a live competent cell, expressing a new phenotype. Griffith’s classic experiment in transformation

Transduction Bacteriophage infect host cells Serve as the carrier of DNA from a donor cell to a recipient cell Generalized Specialized

Genetic transfer based on generalized transduction.

Genetic transfer based on specialized transduction.

Transposon “Jumping genes” Exist in plasmids and chromosomes Contains genes that encode for enzymes that remove and reintegrate the transposon Small transposons are called insertion elements