Microbial Genetics: Structure, Replication, and Gene Function

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Microbial Genetics

Genetics and Genomes

Genetics is the study of inheritance and inheritable traits as expressed in an organism’s genetic material. The genome is the entire genetic complement of an organism, including its genes and nucleotide sequences.

Genome: All genetic material in a cell, including chromosomes and plasmids.
Gene: A segment of DNA that codes for a functional product, usually a protein.
Nucleotide: The basic unit of nucleic acids, composed of a phosphate, pentose sugar, and nitrogenous base.

Structure of nucleic acids and base pairing

Structure of Nucleic Acids

Nucleic acids are polymers of nucleotides. DNA and RNA differ in their sugars and bases. DNA is double-stranded and features complementary base pairing (A-T, G-C), while RNA is usually single-stranded and uses uracil instead of thymine.

Antiparallel: DNA strands run in opposite directions (5’ to 3’ and 3’ to 5’).
Base Pairing: Adenine pairs with thymine (or uracil in RNA), guanine pairs with cytosine.
Backbone: Sugar-phosphate backbone with nitrogenous bases projecting inward.

Structure of nucleic acids and base pairing

Prokaryotic and Eukaryotic Genomes

Prokaryotic genomes typically consist of a single, circular chromosome located in the nucleoid. Eukaryotic genomes are more complex, with multiple linear chromosomes sequestered within a nucleus, and additional DNA in mitochondria and chloroplasts.

Prokaryotes: Haploid, circular chromosomes, may contain plasmids.
Eukaryotes: Diploid, linear chromosomes, chromatin packaging (euchromatin vs. heterochromatin).
Plasmids: Small, circular DNA molecules that replicate independently and may confer advantages (e.g., resistance, virulence).

Prokaryotic genome structure Eukaryotic chromosomal packaging

DNA Replication

Semiconservative Replication

DNA replication is semiconservative: each new DNA molecule consists of one original strand and one newly synthesized strand. The process relies on the complementary structure of DNA.

Key Enzymes: DNA polymerase, helicase, primase, ligase, gyrase, topoisomerase.
Directionality: DNA polymerase synthesizes DNA only in the 5’ to 3’ direction.
Leading Strand: Synthesized continuously toward the replication fork.
Lagging Strand: Synthesized discontinuously away from the fork, forming Okazaki fragments.

Semiconservative model of DNA replication Triphosphate deoxyribonucleotides in DNA synthesis DNA replication process

Okazaki Fragments

Okazaki fragments are short DNA segments synthesized on the lagging strand. They are initiated by RNA primers and later joined by DNA ligase to form a continuous strand.

Reason: DNA polymerase can only add nucleotides in the 5’ to 3’ direction.
Processing: DNA polymerase I replaces RNA primers; DNA ligase seals nicks.

Bidirectional Replication and Methylation

Bacterial DNA replication is bidirectional from a single origin. DNA methylation plays roles in gene expression, replication initiation, protection, and repair.

Bidirectionality of DNA replication in prokaryotes

Gene Function

Genotype and Phenotype

The genotype is the set of genes in the genome, while the phenotype is the physical and functional expression of those genes.

Central Dogma: DNA is transcribed to RNA, which is translated to form polypeptides.

Central dogma of genetics

Transcription

Transcription is the process by which information in DNA is copied as RNA. In prokaryotes, it occurs in the nucleoid; in eukaryotes, in the nucleus, mitochondria, and chloroplasts.

Types of RNA: mRNA, rRNA, tRNA, regulatory RNA, ribozymes, RNA primers.
Steps: Initiation, elongation, termination.
Eukaryotic Processing: Capping, polyadenylation, splicing (removal of introns).

Transcription events in prokaryotes Concurrent RNA transcription Processing eukaryotic mRNA

Translation

Translation is the process in which ribosomes use genetic information in mRNA to synthesize polypeptides. The accuracy of translation depends on correct pairing between mRNA codons and tRNA anticodons.

Participants: mRNA, tRNA, ribosomes, rRNA.
Start Codon: AUG (codes for N-formylmethionine in prokaryotes, methionine in eukaryotes).
Stages: Initiation, elongation, termination.
Ribosome Sites: A (arrival), P (peptide), E (exit).

The genetic code Prokaryotic mRNA molecules lack introns and exons N-formylmethionine vs methionine Transfer RNA structure Ribosomal structures Ribosome and tRNA-binding sites Initiation of translation in prokaryotes Elongation stage of translation Polyribosome in prokaryotes Termination of translation

Regulation of Genetic Expression

Gene expression is regulated to conserve energy. Most genes are expressed at all times, but others are transcribed and translated only when needed. Regulation typically halts transcription or translation.

Operon: A cluster of genes under control of a single promoter and operator.
Inducible Operon: Activated by inducers (e.g., lac operon for lactose catabolism).
Repressible Operon: Continuously transcribed until deactivated by repressors (e.g., trp operon for tryptophan synthesis).

Operon structure CAP-cAMP enhances lac transcription Lac operon, inducible operon trp operon, repressible operon

Regulatory RNA

RNA molecules such as microRNAs (miRNAs), small interfering RNAs (siRNAs), and riboswitches can regulate translation by binding to mRNA or changing shape.

Mutations

Types and Effects of Mutations

A mutation is a change in the nucleotide base sequence of a genome. Most mutations are deleterious, but some may confer advantages. Mutations can be point (affecting one base pair) or gross (involving larger changes).

Point Mutations: Substitutions, insertions, deletions.
Gross Mutations: Inversions, duplications, transpositions.

Types of point mutations and their effects

Mutagens

Mutagens are agents that increase the mutation rate. They include radiation (ionizing and nonionizing) and chemical mutagens (nucleotide analogs, nucleotide-altering chemicals, frameshift mutagens).

Thymine dimer formation by UV light Nucleotide analog structure and effects Action of a frameshift mutagen

DNA Repair Mechanisms

Cells have several methods for repairing damaged DNA, including direct repair, single-strand repair, and error-prone repair (SOS response).

DNA repair mechanisms

Identifying Mutants, Mutagens, and Carcinogens

Mutants are cells that do not repair a mutation. Wild-type cells are those normally found in nature. Methods to recognize mutants include positive selection, negative (indirect) selection, and the Ames test.

Positive selection of mutants Negative selection to isolate tryptophan auxotroph Ames test for mutagenicity

Genetic Recombination and Transfer

Genetic Recombination

Exchange of nucleotide sequences often occurs between homologous sequences, resulting in recombinants—cells with new DNA sequences.

Genetic recombination

Horizontal Gene Transfer in Prokaryotes

Horizontal gene transfer involves the movement of genetic material between cells, not from parent to offspring. Three main types are transformation, transduction, and conjugation.

Transformation: Uptake of DNA from the environment by competent cells.
Transduction: Transfer of DNA via a bacteriophage (virus).
Conjugation: Transfer of DNA requiring cell-to-cell contact, often mediated by pili.

Transformation of Streptococcus pneumoniae Transduction process Bacterial conjugation Conjugation involving Hfr cell

Transposons and Transposition

Transposons are segments of DNA that move from one location to another, causing frameshift insertions. Insertion sequences are the simplest transposons, while complex transposons carry additional genes.

Transposition process Transposons structure

Summary Table: Types of Horizontal Gene Transfer

Type	Mechanism	Key Features
Transformation	Uptake of naked DNA	Requires competence
Transduction	DNA transfer via bacteriophage	Generalized or specialized
Conjugation	Direct cell-to-cell transfer	Requires pili, F plasmid

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