BIOL 340: RNA Processing, Translation, and Prokaryotic Gene Regulation

Comprehensive practice flashcards covering eukaryotic RNA processing (capping, polyadenylation, splicing, editing), prokaryotic translation mechanisms and ribosome structure, and bacterial gene regulation operon dynamics.

What are the three central processing steps that convert eukaryotic pre-mRNA into mature mRNA?

The three central steps are $5'$ capping, $3'$ cleavage and polyadenylation, and splicing.

What chemical structure comprises the eukaryotic $5'$ cap and how is it attached to pre-mRNA?

A 7-methylguanosine ($\text{m}^7\text{G}$) cap is attached to the $5'$ end through a distinctive $5'\text{--}5'$ triphosphate bridge.

What three enzymes act sequentially to install the $5'$ cap on eukaryotic pre-mRNA, and what does each do?

Triphosphatase removes a phosphate from the $5'$ end, guanylyltransferase adds GMP, and methyltransferase transfers a methyl group to form $\text{m}^7\text{G}$.

What are the primary biological functions of the eukaryotic $5'$ cap?

It protects the $5'$ end from exonuclease degradation, and its associated cap-binding proteins assist in nuclear export, splicing, and translation initiation.

What sequence signals specify $3'$ cleavage and polyadenylation in eukaryotic pre-mRNA?

An AAUAAA signal typically positioned $15\text{--}30$ nucleotides upstream of the cleavage site and a downstream sequence element positioned about $20\text{--}40$ nucleotides downstream.

Which enzyme synthesizes the poly(A) tail on pre-mRNA, and what is the typical tail length?

Poly(A) polymerase (PAP) uses ATP to add a non-templated tail of roughly $50\text{--}250$ adenosines.

What is the function of CPSF during eukaryotic mRNA processing?

CPSF helps recognize the cleavage signal and cleave the pre-mRNA at the poly(A) site.

What conserved sequence motifs define splice sites and the branch point in spliceosome-mediated splicing?

A conserved GU at the $5'$ splice site, an internal adenine (A) at the branch point, and an AG at the $3'$ splice site.

What are the two consecutive transesterification reactions that occur during pre-mRNA splicing?

First, the branch-point A attacks the $5'$ splice site, releasing exon 1 and generating an intron lariat. Second, the $3'\text{-OH}$ of exon 1 attacks the $3'$ splice site, ligating the two exons and liberating the lariat intron.

What roles do specific snRNPs play during spliceosome assembly and catalytic activation?

U1 recognizes the $5'$ splice site, U2 binds the branch site exposing the reactive A, and the U4/U5/U6 tri-snRNP joins; activation triggers the release of U1 and U4, allowing U2 and U6 to form the catalytic core while U5 aligns the exons.

What is alternative splicing and what patterns can it follow?

A regulated process where different exon combinations are joined to generate distinct mRNA and protein isoforms from one pre-mRNA; patterns include exon skipping, alternative $5'$ or $3'$ splice sites, and mutually exclusive exons.

Which gene and disease are cited as examples of pathology caused by aberrant RNA splicing?

Duchenne muscular dystrophy, which is linked to splicing errors or disruptions in the dystrophin gene.

How do Group I and Group II self-splicing introns differ in their reaction nucleophiles?

Group I introns use an external guanosine (G) nucleotide to initiate splicing, whereas Group II introns use an internal adenosine (A) residue and form a lariat intermediate.

What is RNA editing and which enzyme carries out A-to-I editing?

RNA editing alters the nucleotide sequence of an RNA molecule post-transcriptionally; A-to-I editing is mediated by ADAR (adenosine deaminase acting on RNA).

How many total codons exist in the genetic code, how many encode amino acids, and what do the remainder do?

There are $64$ codons: $61$ are sense codons specifying amino acids, and $3$ (UAA, UAG, UGA) are nonsense/stop codons that signal termination.

What is meant by the statement that the genetic code is 'degenerate'?

Multiple synonymous codons can encode the exact same amino acid, although each individual sense codon specifies only one amino acid.

How does a tRNA adapter attach to its corresponding amino acid?

An aminoacyl-tRNA synthetase covalently links the correct amino acid to the conserved $3'$ CCA sequence of its cognate tRNA.

What is wobble pairing and how does inosine contribute to it?

Wobble pairing permits nonstandard, relaxed base pairing at the third ($3'$) position of an mRNA codon; an inosine (I) base in the tRNA anticodon allows a single tRNA to pair with multiple synonymous codons.

What are the specific functions of the A, P, and E sites within the ribosome?

The A (aminoacyl) site accepts an incoming aminoacyl-tRNA; the P (peptidyl) site holds the tRNA attached to the growing peptide chain; the E (exit) site retains the deacylated (uncharged) tRNA before its release.

How are the four levels of protein structure defined?

Primary structure is the linear amino acid sequence; secondary structure comprises local hydrogen-bonded motifs ($\alpha$-helices and $\beta$-sheets); tertiary structure is the overall three-dimensional fold of a single polypeptide; quaternary structure is the spatial arrangement of multiple polypeptide subunits.

What sequence interaction correctly positions the bacterial 30S ribosomal subunit over the start codon during translation initiation?

Base pairing between the mRNA Shine–Dalgarno sequence and the complementary sequence at the $3'$ end of the 16S rRNA positions the AUG start codon directly within the ribosomal P site.

What bacterial factors form the 70S initiation complex at the start of translation?

The 30S subunit binds mRNA and positions initiator fMet-tRNA on the AUG codon, followed by the association of the 50S subunit to complete the 70S initiation complex.

What are the functions of EF-Tu and EF-G during the elongation phase of bacterial translation?

EF-Tu–GTP escorts the cognate aminoacyl-tRNA to the ribosomal A site, while EF-G–GTP catalyzes ribosome translocation by advancing the ribosome exactly one codon along the mRNA.

How does translation terminate when a stop codon enters the ribosomal A site?

Protein release factors recognize the stop codon in the A site, promoting hydrolysis and release of the completed polypeptide chain, followed by ribosome dissociation and recycling.

What is the directionality of mRNA translation and polypeptide chain synthesis?

The ribosome translates the mRNA in the $5' \rightarrow 3'$ direction, and the polypeptide chain is elongated from its N-terminus to its C-terminus.

What are the sedimentation coefficients of intact ribosomes and their component subunits in prokaryotes versus eukaryotes?

Prokaryotes contain $70\text{S}$ ribosomes made of $30\text{S}$ and $50\text{S}$ subunits; eukaryotes contain $80\text{S}$ ribosomes made of $40\text{S}$ and $60\text{S}$ subunits.

What is the functional difference between a structural gene and a regulatory gene?

A structural gene codes for an enzyme, functional RNA, or structural protein used in metabolism or cellular architecture; a regulatory gene codes for an RNA or protein that controls the transcription or expression of other genes.

How do promoter and operator DNA elements differ in their function?

A promoter is the sequence recognized and bound by RNA polymerase to initiate transcription; an operator is a distinct DNA sequence where a repressor protein binds to inhibit or prevent transcription.

What distinguishes positive transcriptional regulation from negative transcriptional regulation?

In positive regulation, an activator protein enhances transcription initiation by facilitating RNA polymerase binding; in negative regulation, a repressor protein reduces or blocks transcription initiation by hindering RNA polymerase.

What is an operon in bacterial systems?

A functional unit consisting of a cluster of adjacent structural genes transcribed together into a single polycistronic mRNA, governed by shared cis-regulatory elements such as a promoter and operator.

How do allosteric effectors influence bacterial gene expression?

Allosteric effectors bind reversibly to regulatory proteins (activators or repressors), inducing conformational changes that either promote or inhibit their ability to bind target DNA sequences.