Lectures 5+6: Genomes, Transcription, Translation, and Protein Processing

Genome

all of the genetic information (DNA) that defines an organism, including the chromosome, plasmids and viral DNA

mRNA

messenger RNA: instructions for making proteins

tRNA

transfer RNA: shuttles aa → delivers encoded amino acid to ribosome

rRNA

ribosomal RNA: forms ribsomes, makes protein (translation)

T/F: Genome size does not correlate with lifestyle of an organism. Why?

FALSE.

• Species that colonize diverse niches have larger genomes

• Species with limited niche or those that rely on other species (e.g. intracellular pathogens/symbionts) have smaller genomes – they lose genes over time

Prokaryotic genome differences (from eukaryotic)

• smaller

• lack introns

• less non-coding sequences: greater coding density

• operons: multiple fxn linked genes with a single promoter

  • monocistronic RNA: codes for one protein- one promoter, one gene

  • polycistronic RNA: codes for multiple proteins- ex: one promoter, three genes on one single continuous RNA

Chromosome resides in cytoplasm in a __ state

Supercoiled.

Supercoiled chromosome details

• Reduces volume

• held down by histone-like proteins

• opposite direction to double helix: two strands of negatively coiled DNA are easier to unwind than if they were both positive

Topoisomerases

Enzymes that change DNA supercoiling

• breaking the DNA, passing a section through the break, then resealing the break

DNA replication is conservative/semiconservative?

Semiconservative: DNA replication occurs in both directions, where one parental strand is conserved and inherited by each daughter cell

Plasmids

circular DNA molecules that replicate independently of the bacterial chromosome (also in archaea/eukaryotes)

• Contain nonessential genes that often play critical roles in certain situations (e.g., antibiotic resistance)

• Used to transfer genes in lab

How plasmids replicate (2)

1. Bidirectional replication: starts at single origin and occurs in two directions simultaneously

2. Polling-circle replication: starts at single origin and moves only in one direction

Plasmid tricks to ensure inheritance (3)

1. Carry genes whose functions benefit host microbe under certain conditions (ex: antibiotic resistance)

2. High-copy-number plasmids, making it more likely they’ll be inherited

3. Low-copy-number plasmids evolved dedicated partitioning systems to make sure each daughter cell receives a copy

Secondary chromosome

Genomes with multiple chromosomes are made of a primary chromosome, and one or more smaller secondary chromosomes

• distinguished from plasmids in that they carry at least one essential gene

• evolved from plasmids that captured one or more essential genes from original/primary chromosome

Eukaryotic vs Archaeal chromosomes

Eukartotic: linear, within nucleus, involves mitosis

Archaeal: circular, not in nucleus

In which of the following enzymes would you expect to find a mutation if a bacterial cell shows defects in both chromosome packing and DNA replication?

a) topoisomerase

b) helicase

c) ligase

d) polymerase

A

The genome of a newly-discovered bacterium is comprised of two circular DNA molecules. The larger of the two includes most of the housekeeping genes that are needed for day-to-day activities like metabolism. The genes required for binary fission (cell division) are found on the other molecule. Which of the following phrases provides the most accurate reference for this other molecule, and why?

a) secondary chromosome, because it contains essential genes

b) primary chromosome, because cell division is a critical function

c) plasmid, because it is smaller than the other molecule

d) phagemid, because it likely originated from a replicating virus

A

Noncoding sequences make up a large portion of eukaryotic chromosomes. These include

a) exons.

b) bacteriophages.

c) plasmids.

d) introns.

e) topoisomerases.

D

Archaea resemble eukaryotes in all of the following EXCEPT

a) nuclear membrane.

b) DNA-packing proteins.

c) RNA polymerase.

d) ribosomal components.

e) DNA polymerase.

A

Transcription

Reading a DNA template to make a RNA

Translation

Decoding RNA to make polypeptide

Archaea and eukaryotes share similar transcription and translation machinery and mechanisms compared to those in bacteria; this implies…

Implies an archaeal ancestor that eukaryotes evolved from

RNA polymerase function and composure

Carries out transcription by make RNA copies of a DNA template strand

• Core polymerase: elongation of transcript

• Sigma factor(s): required for initiation of transcript

• Holoenzyme: Core polymerase and sigma factor together

Sigma factor

Helps core polymerase detect the promoter, which signals the beginning of a gene/initiates RNA synthesis

Housekeeping sigma factor

Keeps essential genes and pathways operating, something every cell has

Consensus sequence

Most frequently observed (conserved) sequences

Alterations from consensus sequences reduces promoter strength (less transcription)

In sigma factors with similar consensus sequences, genes with promoter sequence closer to the sigma consensus sequence will be favored for expression

Transcription termination (2)

1. Rho-dependent: relies on protein Rho. Contact btwn Rho and RNA polymerase causes termination

2. Rho-independent: requires region on RNA that forms a hairpin loop and 4-8 consecutive U residues

2 Fundamental criteria that must be met by antibiotics

1. Kill or slow the growth of a pathogen

2. Not harm host (you)

sRNA

Small RNA: regulates transcription or translation

tmRNA

Frees up ribosomes stuck on damaged mRNA

Catalytic RNA

Carries out enzymatic reactions (ex: ribozyme)

Codon

nucleotide triplet

Number of subunits that make up ribosome

Two. Each include rRNA ad numerous proteins

Shine-Dalgarno sequence

Where ribosome binds. Typically 4-8 nucleotides upstream from the start codon

Peptidyltransferase

enzymatic activity where ribsome makes the peptide bonds that stitch aa together

The sigma factor is directly required for

a) transcription initiation.

b) transcription elongation.

c) translation initiation.

d) translation elongation

A

In E. coli, why is sigma-70 considered the “housekeeping” sigma factor?

a) It is expressed in stationary phase to maintain homeostasis.

b) It blocks the random transcription that occurs at low levels

along the chromosome.

c) It keeps essential genes and pathways operating.

d) It binds universally to all E.coli promoters

C

A mutation occurs upstream of gene A in the region encoding the ribosome binding site. Predict how this mutation will impact the levels of mRNA and protein produced for gene A.

a) mRNA will increase, protein will decrease.

b) mRNA will decrease, protein will decrease.

c) mRNA levels will remain constant, protein will increase.

d) mRNA levels will remain constant, protein will decrease.

e) Neither mRNA nor protein levels will change.

D

The termination of transcription can be caused by which of the following?

a) Stem-loop structure

b) Rho factor

c) Stop codon

d) a) and b)

e) a), b), and c)

D

Chaperones are proteins that

a) export other proteins out of the cell.

b) help other proteins fold properly.

c) degrade misfolded proteins.

d) bind other proteins to inactivate them.

B