Microbial Genetics, DNA Structure, and Gene Regulation in Bacteria

What are the three main parts of microbial genetics covered in the notes?

1. Structure, replication, and expression of DNA; 2. Regulation of gene expression; 3. Mechanisms of genetic variation.

What is the objective of studying microbial genetics?

To understand microbial-specific aspects of DNA, gene organization, transcription, and translation.

What is the structure of DNA in archaea?

Circular molecules associated with histones, coiled into nucleosomes.

How does bacterial DNA typically exist?

As closed circular, supercoiled structures associated with basic proteins.

What is the DNA structure in eukaryotes?

Linear molecules associated with histones, coiled into repeating units called nucleosomes.

What is the error frequency during DNA replication in prokaryotes?

10^-9 to 10^-10 per base pair replicated.

What is the replication speed of prokaryotes compared to eukaryotes?

Prokaryotes: 750-1,000 bp/sec; Eukaryotes: 50-100 bp/sec.

What is a replicon?

A unit of DNA that contains an origin of replication and is replicated as a unit.

What is the rolling-circle mechanism in DNA replication?

A rapid replication method used in E. coli conjugation and some small circular genomes, producing many copies from one initiation.

What are DnaA proteins responsible for in DNA replication?

They denature double-stranded DNA.

What is the role of DNA polymerase III?

It is part of the replisome and requires an RNA primer for DNA replication.

What are exons and introns in eukaryotic gene structure?

Exons are coding sequences, while introns are noncoding sequences that interrupt coding sequences.

What is polycistronic mRNA?

mRNA that encodes more than one polypeptide and often has gene products that function together.

What is the structure of prokaryotic promoters?

They contain consensus sequences for RNA polymerase binding, typically at -35 and -10 regions.

What are the three stages of bacterial transcription?

1. Initiation; 2. Elongation; 3. Termination.

What is Rho-independent termination in transcription?

It involves an mRNA hairpin followed by six uridines, leading to the release of RNA polymerase.

What is Rho-dependent termination in transcription?

It requires the Rho factor and does not involve a polyU tail.

What are the unique features of prokaryotic translation?

Polyribosomes and coupled transcription/translation.

What are the components of the prokaryotic ribosome?

70S ribosome composed of 30S and 50S subunits.

What are the stages of translation?

1. Initiation; 2. Elongation; 3. Termination.

What is the role of initiation factors in protein synthesis?

Initiation factors (IF-1, IF-2, IF-3) ensure proper initiation of protein synthesis.

What are the three phases of elongation in protein synthesis?

The three phases are aminoacyl-tRNA binding, transpeptidation reaction, and translocation.

What are the ribosome tRNA binding sites?

The ribosome has three tRNA binding sites: A (aminoacyl), P (peptidyl), and E (exit).

What happens during translocation in protein synthesis?

Peptidyl-tRNA moves from the A site to the P site, the ribosome moves down one codon, and empty tRNA leaves the P site.

What are release factors (RFs) and their role in protein synthesis?

Release factors aid in the recognition of stop codons during termination of protein synthesis.

What are the three stop codons in protein synthesis?

The stop codons are UAA, UAG, and UGA.

What is the process of splicing in protein processing?

Splicing involves the removal of part of the polypeptide before folding, with inteins being removed portions and exteins being the remaining portions.

What is the function of molecular chaperones in protein folding?

Molecular chaperones aid in the folding of nascent polypeptides and protect cells from thermal damage.

What are some examples of molecular chaperones in E. coli?

Examples include DnaK, DnaJ, GrpE, GroEL, and GroES.

How do protein folding processes differ between prokaryotes and eukaryotes?

In prokaryotes, folding occurs after the entire polypeptide is synthesized, while in eukaryotes, domains fold independently immediately after synthesis.

What triggers the upregulation of heat shock proteins?

Heat shock proteins are upregulated when E. coli experiences a temperature increase from 30°C to 42°C.

What is the Sec-dependent pathway in protein translocation?

The Sec-dependent pathway translocates unfolded proteins from the cytoplasm across or into the plasma membrane.

What is the role of the Tat pathway in protein translocation?

The Tat pathway translocates fully-folded proteins across the inner membrane.

What characterizes the Type I protein secretion pathway?

Type I secretion pathway transports proteins directly from the cytoplasm across both membranes in a one-step process.

What is the function of the Type IV secretion pathway?

The Type IV pathway is involved in the secretion of protein and DNA, utilizing both Sec-dependent and independent mechanisms.

What is the significance of the Type II secretion pathway?

Type II pathway transports proteins from the periplasm across the outer membrane, often involving toxins and proteases.

What is the Type III protein secretion pathway known for?

Type III pathway secretes proteins from the cytoplasm across both membranes and into host cells, often associated with virulence factors.

What is the general secretion pathway in prokaryotes?

The general secretion pathway translocates unfolded proteins from the cytoplasm across or into the plasma membrane.

What is the role of chaperone proteins during protein translocation?

Chaperone proteins keep preproteins unfolded during translocation.

What is the Sec translocon?

The Sec translocon is a membrane channel that facilitates the transfer of proteins and removes the signal peptide during translocation.

What is the primary objective of studying microbial genetics?

To gain an understanding of the mechanisms by which bacteria regulate gene expression.

Why do bacteria regulate gene expression?

To conserve energy and raw materials, adapt to environmental changes, and maintain homeostasis.

What are the three categories of gene expression regulation?

Constitutive, inducible, and repressible.

What is positive transcriptional control?

It allows initiation of transcription through activator proteins.

What is negative transcriptional control?

It inhibits initiation of transcription through repressor proteins.

What happens to the lac operon in the presence of lactose?

Lactose inactivates the LacI repressor, allowing transcription to occur.

What is the role of the LacI protein in the lac operon?

LacI acts as a repressor that binds to the operator to inhibit transcription when lactose is absent.

What is the function of the trp operon?

It regulates the synthesis of tryptophan and is repressed when tryptophan is abundant.

What is catabolite repression?

A regulatory mechanism where the presence of a preferred carbon source (like glucose) inhibits the expression of other catabolic operons.

What is the role of catabolite activator protein (CAP) in gene regulation?

CAP binds to the CAP binding site to facilitate RNA polymerase binding, promoting transcription when cAMP levels are high.

What is attenuation in transcriptional regulation?

It is the halting of transcription elongation prior to termination based on metabolite availability.

What are alternate sigma factors?

Proteins that direct RNA polymerase to different promoters, allowing differential gene expression.

What is a regulon?

A group of operons controlled by the same regulatory protein, often associated with a common function.

What is a modulon?

A regulon that includes operons controlled separately, such as in catabolite repression.

What is the significance of quorum sensing in bacteria?

It allows bacteria to communicate and coordinate behavior based on population density, often regulating bioluminescence.

What is the function of the response regulator in a two-component regulatory system?

It binds to DNA to control transcription in response to signals detected by the sensor kinase.

What is the role of the sensor kinase in bacterial signal transduction?

It detects external changes and transmits signals to the response regulator, often through phosphorylation.

What is diauxic growth?

A biphasic growth pattern observed in bacteria when two different carbon sources are available, utilizing one before the other.

What happens to the trp repressor when tryptophan levels are low?

The trp repressor is inactive, allowing transcription of the trp operon to proceed.

What is the role of allolactose in the lac operon?

Allolactose binds to LacI, inactivating it and allowing transcription of the operon.

What is the effect of glucose on cAMP levels?

When glucose is present, cAMP levels are low, inhibiting the activation of CAP and reducing transcription of certain operons.

What is the relationship between synthesis and degradation in gene expression regulation?

Gene expression is based on maintaining a balance between the synthesis of enzymes and their degradation.

What is the function of methyl-accepting chemotaxis proteins (MCP) in E. coli?

MCPs bind chemoattractants and regulate flagellar rotation to direct movement toward favorable environments.

What is the role of CheY in E. coli chemotaxis?

CheY, when phosphorylated, interacts with FliM to induce clockwise rotation of the flagella, leading to tumbling.

What is the significance of the LuxR protein in V. fischeri?

LuxR acts as a transcriptional activator that requires AHL to activate the expression of bioluminescence genes.

What is the function of riboswitches in transcriptional regulation?

Riboswitches are RNA elements that change conformation in response to metabolite binding, affecting RNA polymerase activity.

What is a mutation?

A stable, heritable change in nucleotide sequence that may or may not affect phenotype.

What are the two main types of mutations?

Spontaneous mutations and induced mutations.

What causes spontaneous mutations?

Errors in DNA replication, DNA damage, or transposon insertion.

What are tautomeric shifts?

Alterations in hydrogen bonding of nucleotides that can lead to replication errors.

What are frameshift mutations?

Mutations caused by the deletion or addition of base pairs that alter the reading frame.

What are induced mutations?

Mutations caused by chemical or physical agents that damage or alter DNA.

Give an example of a base analog.

5-bromouracil, which is a thymine analog that can bind to guanine or adenine.

What are intercalating agents?

Chemical agents like ethidium bromide that insert themselves between DNA bases, causing mutations.

What is the difference between forward mutations and reversion mutations?

Forward mutations change wild type to mutant form, while reversion mutations change mutant phenotype back to wild type.

What are silent mutations?

Mutations that do not change the amino acid sequence of a protein.

What are auxotrophs?

Mutants that cannot synthesize a product required for growth, needing supplements.

What is horizontal gene transfer?

The transfer of DNA from a donor organism to a recipient, enhancing genetic variability.

What are the three mechanisms of horizontal gene transfer?

Conjugation, transformation, and transduction.

What is homologous recombination?

A process where DNA strands break and reunite, resulting in new nucleotide sequences.

What are transposable elements?

Mobile DNA segments that can move within the genome and do not require large homology regions.

What are insertion sequences?

The simplest type of transposable elements that encode only the enzymes required for transposition.

What are composite transposons?

Transposable elements that carry additional genes, such as those for antibiotic resistance.

Describe the 'cut-and-paste' method of transposition.

Transposase cuts the transposable element and inserts it into a new target site, generating direct repeats.

What is the replicative method of transposition?

Transposase makes cuts, and a copy of the transposon is inserted at a new site while the original remains.

What is recombination in genetics?

Rearranging or combining of one or more nucleic acid molecules to produce a new nucleotide sequence.

What are the types of recombination?

1. Homologous recombination 2. Site-specific recombination 3. Transposition.

What is site-specific recombination?

Insertion of primarily nonhomologous DNA into a chromosome, involving only a short region of homology, often during viral genome integration.

What is transposition?

Movement of segments of DNA within the genome, sometimes referred to as 'jumping genes' or transposons.

What are the two methods of transposition?

1. Simple or 'cut-and-paste' 2. Replicative.

What occurs during simple transposition?

Transposase recognizes the ends of the transposable element, cuts them, cleaves the new target site, and ligates them, generating direct repeats of flanking host DNA.

What happens during replicative transposition?

Transposase makes cuts, strands are exchanged, and resolvase catalyzes recombination, leaving the original transposon at the parental site and inserting a copy at the target site.

What are the effects of transposition?

Can cause mutations in coding regions, arrest of translation or transcription, activation of genes, generation of new plasmids, and transfer of resistance among plasmids/chromosomes.

What are bacterial plasmids?

Small, circular double-stranded DNA molecules that are replicons and can exist as episomes.

What are fertility (F) factors?

Conjugative plasmids that play a major role in conjugation, such as the F factor of E. coli.

What are resistance (R) factors?

Plasmids that carry antibiotic resistance genes, which can destroy or modify antibiotics.

What are col plasmids?

Plasmids that encode colicin, a type of bacteriocin that destroys closely-related bacteria.

What are virulence plasmids?

Plasmids that carry genes conferring resistance to host defense mechanisms and may encode toxins.

What is bacterial conjugation?

The transfer of DNA by direct cell-cell contact, initially discovered by Joshua Lederberg and Edward Tatum.

What was Bernard Davis' U-tube experiment?

An experiment demonstrating that contact is required for bacterial conjugation, as no prototrophs were obtained when contact was prevented.