Microbiology: A Systems Approach PowerPoint to accompany Microbiology: A Systems Approach Cowan/Talaro Chapter 9 Microbial Genetics Copyright The McGraw-Hill Companies, Inc. Permission required for reproduction or display.

Chapter 9 Topics - Genetics - Flow of Genetics - Regulation - Mutation - Recombination

Genetics Genome Chromosome Gene Protein Genotype Phenotype

The sum total of genetic material of a cell is referred to as the genome. Fig. 9.2 The general location and forms of the genome

Chromosome Procaryotic Eucaryotic Histonelike proteins condense DNA Eucaryotic Histone proteins condense DNA Subdivided into basic informational packets called genes

Genes Three categories Genotype Phenotype Structural – Code for proteins or peptides Regulatory Encode for RNA Genotype The actual gene types or genetic makeup Phenotype Expression of the genotypes ( measurable or observable)

Flow of Genetics Replication “Central Dogma” DNA =>DNA “Central Dogma” DNA =>mRNA=>Protein Transcription – DNA copied into RNA Translation – mRNA used to build a protein or peptide chain

Representation of the flow of genetic information. Fig. 9.9 Summary of the flow of genetic information in cell.

DNA Structure Replication

DNA is lengthy and occupies a small part of the cell by coiling up into a smaller package. Fig. 9.3 An Escherichia coli cell disrupted to release its DNA molecule.

Structure Nucleic Acid composed of nucleotides Nucleotide Phosphate 5 carbon sugar (Deoxyribose) Nitrogenous base Double stranded helix Each strand has carbon-phosphate covalently linked to form the “backbone” Nitrogenous bases stick out towards the middle Right & left side are linked by H-bonds formed between the bases in the middle ( Complementary base pair bonding) Every 10 nucleotides, the DNA twists – forms the helix Antiparallel arrangement – strands have a 5’ end and a 3’ end. The two strands run in opposite directions.

Nitrogenous bases Purines Pyrimidines Adenine Guanine Thymine Cytosine Uracil (only found in RNA – replaces thymine)

Purines and pyrimidines pair (A-T or G-C) and the sugars (backbone) are linked by a phosphate. Fig. 9.4 Three views of DNA structure

DNA Structure

Replication Semiconservative Enzymes Leading strand Lagging strand Okazaki fragments

Semiconservative Replication New strands are synthesized in 5’ to 3’ direction Each DNA strand acts as a template to recreate the opposite (complementary) strand. When the two new DNA strands are formed, each has 1 original strand ( the parent or template strand) and one newly synthesized ( daughter) strand

Semiconservative replication of DNA synthesizes a new strand of DNA from a template strand. Fig. 9.5 Simplified steps to show the semiconservative replication of DNA

Enzymes Topoisomerases (Gyrase) – relieves supercoiling Helicase – “unzips” – separates the two DNA strands DNA polymerase III – Adds bases to the new DNA chain; proofreads for mistakes DNA polymerase I – Removes the primer; closes gaps; repairs mismatches Primase – makes a short chain of nucleic acids called a “primer” Ligase – seals the Okazki fragments together; final binding of nicks in DNA during synthesis and repair

The function of important enzymes involved in DNA replication. Table 9.1 Some enzymes involved in DNA replication

Leading strand DNA polymerase can only add bases going in the 5’ to 3’ direction RNA primer initiates the 5’ to 3’ synthesis of DNA in continuous manner ( the new DNA is made in one continuous strand)

Lagging strand On the lagging strand, the new area being exposed for copying is running in the opposite direction of the leading strand ( 3’ to 5’) DNA pol can only add bases in the 5’ 3’ direction Multiple primers are laid down Short pieces of DNA called Okazaki fragments are synthesized Okazaki fragments are ligated together to form one continuous strand (DNA ligase)

The steps associated with the DNA replication process. Fig. 9.6 The bacterial replicon: a model for DNA Synthesis

Refinements and Details of Replication

RNA Transcription – the synthesis of RNA from a DNA template synthesize Messenger RNA (mRNA) Transfer RNA (tRNA) Ribosomal RNA (rRNA) Ribozymes and spliceosomes (catalytic RNA) Regulatory RNA Codon – set of 3 RNA bases that code for an amino acid

mRNA Copy of a structural gene or genes of DNA Can encode for multiple proteins on one message Thymidine is replaced by uracil Triplet Code -- the message is organized into codons (1 codon= three bases  codes for 1 amino acid)

Transcription In a gene there are 2 strands of DNA – the coding strand which contains the information sequence to build the protein and the opposite, complementary template strand. In transcription, only 1 strand of DNA is copied, not both. The coding strand has the information to build the protein but it is not copied directly. Copying a nucleic acid occurs by reading the template strand and synthesizing a complementary strand. This new mRNA strand will have the same information as the coding strand of the DNA

Transcription RNA polymerase catalyzes the reaction -- reads the template strand and makes a complementary strand of mRNA. Synthesis occurs in 5’ to 3’ direction You don’t copy the entire strand of DNA – only the sequence that is the gene. So you need to know where to start copying and where to end. The promoter region is a short sequence of bases that occurs right before the start of the gene and signals to RNA pol where to start copying. Termination sequences occur at the end of the gene and signal to RNA pol to stop copying.

3 Stages of Transcription Initiation - RNA pol binds to the promoter region on the template strand. Promoter regions contain typical repeat sequences that are recognized as the start of a gene. The promoter region itself does not get copied. Elongation – The RNA pol moves down the template strand of DNA in the 3’ 5’ direction, unwinding the DNA. It reads the strand and builds the complementary RNA strand in the 5’  3’ direction. The section behind the RNA pol that has already been copied winds back together. Termination – When the termination sequences are reached, the RNA pol falls of the DNA and the mRNA transcript is released.

The synthesis of mRNA from DNA. Fig. 9.12 The major events in mRNA synthesis

Transcription

Transcription

Transcription

tRNA Complimentary sequences form hairpin loops Amino acid attachment site Anticodon – complementary to the codon; makes the tRNA specific to carry only 1 amino acid. Participates in translation (protein synthesis)

Important structural characteristics for tRNA and mRNA. Fig. 9.11 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. Fig. 9.9 Summary of the flow of genetics

Codons Triplet code – A set of 3 mRNA bases that specifies a given amino acid The genetic code is redundant. There are 64 codons but only 20 amino acids: some amino acids - more than one codon can specify the same amino acid. 64 codons 61 code for amino acids Start codon – AUG is the start codon and it also codes for methionine 3 Stop codons (Nonsense codons) – do not code for any amino acids but stop translation

The codons from mRNA specify a given amino acid. Fig. 9.14 The Genetic code

Genetic Code Redundancy: Certain amino acids are represented by multiple codons Allows for the insertion of correct amino acids even when mistakes occur in the DNA sequence Wobble: Only the first two nucleotides are required to encode the correct amino acid The third nucleotide does not change its sense Permits some variation or mutation without altering the message Universal The genetic code is translated almost identically in all organisms

Representation of the codons and their corresponding amino acids. Fig. 9.15 Interpreting the DNA code

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

Participants involved in the translation process. Fig. 9.13 The “players” in translation

Translation Ribosomes bind mRNA at the 5’ end, near the start codon (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. Fig. 9.16 The events in protein synthesis

For procaryotes, translation can occur at multiple sites on the mRNA while the message is still being transcribed. Fig. 9.17 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 Transcription and translation are not simultaneous Pre-mRNA is modified Introns – (non-coding sequences) removed Exons – (coding sequences) spliced together 5’ Guanine cap added ( helps to place the ribosome on the 5’ end of the mRNA) Poly-A tail added at 3’ end helps keep the mRNA from being degraded by enzymes in the cytoplasm

The processing of pre-mRNA into mRNA involves the removal of introns. Fig. 9.18 The split gene of eucaryotes

Proteins are modified after translation Posttranslational modifications: Proteins begin to fold upon themselves to achieve their tertiary conformation even before the peptide chain is released methionine may be clipped off Cofactors may be added Some join with other proteins to form a quaternary structure