Bacterial Genetics and Molecular Biology Practice Flashcards

Comprehensive practice flashcards covering bacterial DNA replication, transcription, translation, protein folding, cloning, and the human microbiome based on lecture transcripts.

Nucleotide

The monomer building block of DNA consisting of a deoxyribose sugar, a negatively charged phosphate group, and a nitrogenous base.

Phosphodiester bond

The covalent chemical bond that links nucleotides together to form the backbone of a DNA strand.

Plectonemes

Branched loops formed when bacterial DNA supercoils to heavily condense the genome.

Nucleoid Associated Proteins (NAPs)

DNA binding proteins that organize genomic DNA into a nucleoid in bacteria, substituting for eukaryotic histones.

Origin of replication (ori)

The specific single point on a bacterial chromosome where replication begins.

Termination site (ter)

The end-point for DNA replication located at the opposite pole of the cell from the origin.

DNA Gyrase

A protein also known as Isomerase that functions to unwind DNA supercoils during replication.

Helicase

The enzyme responsible for unzipping the DNA double helix to create single strands for replication.

Single-stranded DNA binding proteins (SSB)

Proteins that stabilize and keep the separated DNA strands apart during the replication process.

DNA primase

The enzyme that synthesizes the RNA primer required to start DNA synthesis.

DNA polymerase III

The enzyme that binds to single-stranded DNA and adds nucleotides in the $5' \rightarrow 3'$ direction.

DNA ligase

The enzyme that joins Okazaki fragments or newly formed DNA fragments together by repairing phosphodiester bonds.

Catenane

Interlinked circular DNA molecules formed after replication that must be resolved into two separate chromosomes.

FtsK-XerCD

The recombination machinery that resolves chromosome dimers at the ter sites by cleaving Holliday junctions.

H-NS (Histone-like nucleoid structuring protein)

A NAP that preferentially binds curved DNA, covers 1% of the chromosome, and packs side-by-side between DNA strands.

FIS (Factor for inversion stimulation)

A sequence-specific DNA binding protein that bends DNA $50-90$ degrees and acts as a transcriptional activator.

Transcription factories

Specialized sites formed when multiple RNA polymerase molecules and active gene regions cluster together inside the nucleus.

Exponential phase chromosome state

The growth phase where the bacterial chromosome becomes densely packed.

Stationary phase chromosome state

The growth phase where the bacterial chromosome becomes relaxed and expands.

Open Reading Frame (ORF)

The portion of a gene that contains the sequence of bases that could potentially encode a protein.

Promoter

A specific DNA region where RNA polymerase binds to initiate transcription.

Messenger RNA (mRNA)

A type of RNA that carries genetic code instructions for the creation of new proteins.

Transfer RNA (tRNA)

Small adaptor molecules (~80 nucleotides) that decode mRNA into proteins and align amino acids.

Ribosomal RNA (rRNA)

The RNA components that serve as subunits of ribosomes to synthesize proteins.

Coding strand

The DNA strand used as a sequence reference where T is replaced with U in the resulting RNA transcript.

Template strand

The DNA strand read by RNA polymerase in the $3' \rightarrow 5'$ direction during transcription.

RNA polymerase Core Enzyme

The bacterial transcription complex consisting of two $\alpha$ subunits, one $\beta$ subunit, and one $\beta'$ subunit.

Sigma factor ($\sigma$)

An accessory subunit that bind reversibly to RNA polymerase to form the holoenzyme and recognize promoters.

Transcription start point (+1)

The exact location on the DNA where the synthesis of mRNA begins.

Hairpin loop

An RNA secondary structure formed at the self-complementary termination sequence that causes RNA-poly to dissociate.

Shine Dalgarno sequence

A ribosomal binding site in bacterial mRNA that helps initiate translation of a protein.

Start Codon

The specific triplet sequence AUG that signals the beginning of translation.

Stop Codon

Triplet sequences (UAG, UAA, UGA) that signal the termination of translation.

Inosine

A modified adenine base found in tRNA as a result of post-transcriptional modifications.

Aminoacyl-tRNA synthetases

Enzymes that determine the attachment of the correct amino acids to tRNA based on its 3D shape.

30S subunit

The small ribosomal subunit in prokaryotes that contains 16S rRNA and binds to the Shine Dalgarno site.

50S subunit

The large ribosomal subunit in prokaryotes containing 5S and 23S rRNA and peptidyltransferase activity.

Peptide bond

A covalent bond formed between two amino acids via a condensation reaction.

Degenerate code

The property of the genetic code where multiple codons (64 total) can encode the same amino acid (20 total).

Wobble base pairing

A mechanism that allows a single tRNA to recognize multiple codons by flexible pairing at the third base position.

A site (aminoacyl-tRNA)

The ribosomal binding site where a new tRNA enters the ribosomal complex.

P site (peptidyl-tRNA)

The ribosomal binding site where the tRNA attaches to the growing polypeptide chain.

E site (exit)

The ribosomal binding site where deacetylated tRNA leaves the complex.

Peptidyltransferase

The activity within the 50S subunit that catalyzes the covalent bond formation between amino acids.

Release factors (RF)

Proteins that fill the A site of the ribosome when a stop codon is reached to trigger polypeptide release.

Cistron

A segment of DNA that codes for a specific protein or polypeptide.

Polycistron

A single mRNA molecule that contains codes for more than one protein, common in bacteria.

Repressor

A regulatory protein that binds to DNA to prevent the transcription of a gene.

Activator

A regulatory protein that stabilizes the interaction of RNA-poly with the promoter to induce transcription.

Inducer

A small ligand molecule that binds to a regulatory protein to enable or disable its binding to DNA.

DNA binding domain (DBD)

The modular part of a regulatory protein that physically attaches to the DNA operator.

Effector binding domain (EBD)

The modular part of a regulatory protein where a ligand binds to trigger a conformational change in the DBD.

Histidine Kinase (HK)

A sensory protein in the inner membrane that phosphorylates itself on a His residue upon sensing environmental cues.

Response Regulator

A cytoplasmic protein that receives a phosphate group from Histidine Kinase on an aspartate residue to act as a transcription factor.

AraC

A regulatory protein for the arabinose operon that acts as a repressor when elongated and an activator when compact.

Attenuation

A control mechanism where transcription is stopped prematurely before the RNA polymerase reaches the structural gene.

Leader sequence (L)

The first 140 base pairs of mRNA transcribed upstream of the start codon, critical for attenuation.

rpoH mRNA (thermosensitive riboswitch)

An mRNA whose secondary structure acts as a thermostat, occluding the Shine Dalgarno sequence at normal temperatures.

sRNA

Short, non-coding RNA molecules that regulate expression by binding target mRNAs to trigger breakdown or stop translation.

Slipped strand mispairing

An occurrence during DNA replication where repetitive sequences misalign, leading to insertions or deletions that switch gene activity.

PhoPQ system

A two-component system in bacteria that responds specifically to low magnesium levels.

PmrAB system

A two-component system in bacteria that responds specifically to low iron levels.

PCR (Polymerase Chain Reaction)

A laboratory technique used to amplify specific segments of DNA using cycling temperatures.

Taq polymerase

A heat-stable DNA polymerase used in PCR that remains functional up to $95^{\circ}\text{C}$.

Denaturation (PCR)

The first step of PCR ($94-98^{\circ}\text{C}$) where double-stranded DNA template is melted into single strands.

Annealing (PCR)

The second step of PCR ($50-65^{\circ}\text{C}$) where primers attach to their complementary sequences on the template.

Extension (PCR)

The final step of PCR ($72^{\circ}\text{C}$) where DNA polymerase builds new strands from the primers.

ddNTP (dideoxynucleoside triphosphate)

Nucleotides labeled with chromophores used in DNA sequencing to terminate chain elongation.

Restriction Enzyme

An enzyme that cleaves phosphodiester bonds at specific palindromic sites in the DNA duplex.

DNA Palindrome

A nucleic acid sequence where the $5' \rightarrow 3'$ reading matches on both complementary strands.

Sticky ends

DNA fragments produced by restriction enzymes that have single-stranded overhangs.

Blunt ends

DNA fragments produced by restriction enzymes that are flush and even with no overhanging bases.

Isoschizomer

A pair of restriction enzymes from different sources that recognize and cleave the exact same nucleotide sequence.

Agarose

A polymer of repeating di-galactan units used to create porous gels for separating DNA by size.

Cloning vector

A small plasmid containing a selectable marker, multiple restriction sites, and an origin of replication.

Alkaline Phosphatase (AP)

An enzyme used to remove $5'$ phosphates from cut DNA to prevent the re-ligation of empty vectors.

Klenow fragment

A fragment of DNA-pol1 used to fill in $5'$ overhangs or remove $3'$ overhangs to create compatible DNA ends.

Bacterial transformation

The process of making cells permeable (e.g., using $CaCl_2$ and heat shock) to take up foreign plasmid DNA.

Insertional inactivation

A technique where foreign DNA is placed inside a marker gene, breaking its function to identify recombinant clones.

Blue-white screening

A screening method where successful DNA insertion is indicated by white colonies, while empty plasmids produce blue colonies.

X-gal

A chromogenic substrate that turns blue when cleaved by the enzyme $\beta$-galactosidase.

IPTG

A gratuitous inducer that activates the lac operon to express $\beta$-galactosidase without being metabolized.

$\alpha$-complementation

The process where two inactive fragments of $\beta$-galactosidase associate to restore enzymatic activity.

JM109 strain

An E. coli strain with a lacZ deletion (lacZ$\Delta$M15) used for $\alpha$-complementation in blue-white screening.

Colony hybridization

A molecular technique using labeled nucleotide probes to identify specific bacterial clones on a membrane.

Hydrophobic collapse

The stage of protein folding where the protein snaps into a compact globular structure to hide non-polar regions.

Trigger Factor (TF)

A ribosome-associated chaperone (PPIase) that facilitates the isomerization of proline during co-translational folding.

ClpB

A high-activity disaggregase that uses ATP to pull polypeptide chains through a channel to disentangle protein clumps.

DnaK/DnaJ/GrpE system

A chaperone system that spots exposed hydrophobic areas on aggregates and recruits ClpB for disaggregation.

GroEL/ES Chaperonin

A macromolecular machine consisting of a nano-cage that provides a safe space for proteins to refold using ATP.

Integral membrane proteins

Proteins with hydrophobic domains (often $\alpha$-helices) that span or sit inside the phospholipid bilayer.

Signal recognition particle (SRP) pathway

A pathway using Ffh and sRNA to transport proteins destined for the inner membrane or Sec system.

Sec-dependent pathway

A system where SecA uses ATP to push unfolded proteins through the SecYEG channel into the periplasm.

LepB (signal peptidase)

The enzyme that removes the cleavable signal sequence from proteins once they reach the periplasm.

Twin Arginine Transport (TAT) pathway

A pathway that uses proton motive force (pmf) to transport fully folded proteins across the inner membrane.

Oxidoreductases

Enzymes like DsbA and DsbC that form or fix vital disulfide bonds during protein folding in the cell envelope.

BAM complex

The machinery responsible for flipping unfolded proteins into the outer membrane of Gram-negative bacteria.

ClpP

An ATP-dependent endoprotease with a proteolytic core consisting of two rings used for protein degradation.

Fusion vector

A plasmid designed to produce a translational fusion with an N-terminal partner protein to aid in purification.

LacY

The gene product in the lactose operon responsible for importing lactose into the bacterial cell.