Chapter Summary

8.1

  • RNA polymerase holoenzyme consisting of core RNA polymerase and a sigma factor executes transcription of a DNA template strand.
  • Sigma factors help core RNA polymerase locate consensus promoter sequences near the beginning of a gene. The sequences identified and bound by E. coli sigma-70 are located –10 and –35 bp upstream of the transcription start site.
  • Dynamic changes in gene expression result when the relative levels of different sigma factors change.

8.2

  • A closed complex occurs on initial binding of RNA polymerase to promoter DNA.
  • Opening (melting) of the DNA strands produces a “bubble” of DNA around the polymerase and forms the open complex.
  • Rho-dependent and Rho-independent mechanisms mediate transcription termination.
  • Rifamycin B is an antibiotic that selectively inhibits transcription initiation of bacterial RNA polymerase.
  • Actinomycin D is an antibiotic that nonselectively inhibits transcription elongation.
  • Messenger RNA molecules encode proteins.
  • Ribosomal RNA are stable molecules found in ribosomes and contain unusual modified bases.
  • Transfer RNA are stable molecules that bind amino acids and contain modified bases.
  • Small RNA molecules can regulate gene expression, have catalytic activity (catalytic RNA), or function as a combination of tRNA and mRNA (tmRNA).
  • Half-lives of RNA species vary within the cell.

8.3

  • Triplet nucleotide codons in mRNA encode specific amino acids. Transfer RNA molecules interpret the genetic code and bring specific amino acids to the A (acceptor) site in the ribosome.
  • Specific codons mark the beginning and end of a gene, but the Shine-Dalgarno sequence in mRNA located before the start codon helps the ribosome find the correct reading frame in the mRNA.
  • Initiation of protein synthesis in bacteria requires three initiation factors that bring the ribosomal subunits together on an mRNA molecule.
  • Peptidyltransferase activity of the ribosome is carried out by ribosomal RNA, not protein. Elongation of translation occurs when the ribosome ratchets one codon length along the mRNA.
  • Translation terminates upon reaching a stop codon. The ribosome pauses because it cannot find an appropriate tRNA. A release factor enters the A site and triggers peptidyltransferase activity, thus freeing the completed protein from tRNA in the A site.
  • Ribosome release factor and EF-G bind to the A site to dissociate the two ribosomal subunits from the mRNA.
  • Antibiotics that affect translation can prevent 70S ribosome formation (streptomycin), inhibit aminoacyl-tRNA binding to the A site (tetracycline), interfere with peptidyltransferase (chloramphenicol), trigger peptidyltransferase prematurely (puromycin), cause abortive translocation (erythromycin), or prevent translocation (fusidic acid).
  • Transcription and translation in prokaryotes are coupled.
  • RNA polymerase pauses during transcription to allow the slower-translating ribosomes to stay close. This pause minimizes exposure of mRNA to degradative cellular enzymes.
  • tmRNA rescues ribosomes stuck on damaged mRNA that lacks a stop codon.

8.4

  • Protein modifications are made after translation.
  • The N-terminal amino acid (N-formylmethionine) can be removed by methionine deformylase.
  • An inactive precursor protein can be cleaved into a smaller active protein, or other groups can be added to the protein (for example, phosphate or AMP).
  • Chaperone proteins help translated proteins fold properly.

8.5

  • Special protein export mechanisms are used to move proteins to the inner membrane, the periplasm, the outer membrane, and the extracellular surroundings.
  • N-terminal amino acid signal sequences help target membrane proteins to the membrane.
  • The general secretory system involving the SecYEG translocon can move unfolded proteins to the inner membrane or periplasm.
  • The signal recognition particle (SRP) pauses translation of a subset of proteins that will be placed into the membrane.
  • SecB protein binds to certain unfolded proteins that will eventually end up in the periplasm, and pilots them to the SecYEG translocon.
  • The twin arginine translocase (TAT) can move a subset of already folded proteins across the inner membrane and into the periplasm.
  • Type I secretion systems are ATP-binding cassette (ABC) mechanisms that move certain secreted proteins directly from the cytoplasm to the extracellular environment.

8.6

  • All proteins are eventually degraded.
  • The N-terminal rule describes one type of degradation signal (degron) that marks the half-life of a protein (how long it takes 50% of the protein to degrade).
  • ATP-dependent proteases such as Lon or ClpP usually initiate degradation of a large protein.
  • Damaged proteins randomly enter chaperone-based refolding pathways or degradation pathways until the protein is repaired or destroyed.

8.7

  • Annotation requires computers that look for patterns in DNA sequences. Annotation predicts regulators, ORFs, rDNA, and tRNA. Similarities in protein sequence (deduced from the DNA sequence) are used to predict protein structure and function.
  • An open reading frame (ORF) is a sequence of DNA predicted by various sequence cues to encode an actual protein.
  • Eukaryotic genes contain introns and exons, making computer predictions of an ORF more difficult than with prokaryotic genes.
  • DNA alignments of similar genes or proteins can reveal evolutionary relationships.
  • Paralogs and orthologs arise from gene duplications. Paralogous genes coexist in the same genome but have different functions. Orthologous genes occur in the genomes of different species but produce proteins with similar functions.