Bacterial Genomes and Central Processes

Question 1

What does the bacterial genome consist of?

Answer 1

The single covalently circular duplex molecule - bacterial chromosome

Plasmids

99% is unique DNA

Question 2

What are single nucleotide variants?

Answer 2

Single nucleotide changes that occur over generations

Can be used in the identification of pathogenic subspecies or strains

Question 3

What are pathogenicity Islands?

Answer 3

Cluster of genes encoding proteins involved in pathogenesis (adhesions, invasions, exotoxins and secretory pathways)

Same island can be present in different species possibly because it is a transposon

GC content of island is often different than the host genomes

Question 4

What is semi-conservative replication?

Answer 4

each daughter duplex DNA molecule is composed of one parental strand and one newly-synthesized strand

Question 5

What is the origin of replication (OriC)?

Answer 5

Single site on chromosome where all DNA replication is initiated

Question 6

Describe bacterial DNA polymerases.

Answer 6

Pol I, II, and III are required for DNA synthesis

All share 5’ to 3’ polymerase activity

All share 3’ to 5’ exonuclease (proofreading) activity necessary for maintaining a lower error rate

Question 7

Describe bacterial supercoiling.

Answer 7

Bacterial chromosomes have negative superhelicity which means that it can readily convert to a molecule with unwound helical regions

Absolutely necessary for DNA replication, and is the target of some antibiotics (e.g. quinolones)

Question 8

What are the three steps in bacterial chromosome replication during cell division?

Answer 8

Chromosome is attached to cytoplasmic membrane protein complex

Initiation of chromosome replication coincides with septum formation

As cell wall growth elongates, chromosomes are pulled into daughter cells

Question 9

What are the four types of RNA molecules in a cell?

Answer 9

rRNA

tRNA

mRNA

microRNA

Question 10

What is DNA-dependent RNA polymerase (RNAP)?

Answer 10

Single enzyme that transcribes all four types of RNA

Question 11

What are two alternative subunits for the RNAP holoenzyme?

Answer 11

Sigma factor that imparts initiation specificity to the holoenzyme - recognizes -10 and -35 positions of the promoter

Rho factor that assists in termination in some genes; other genes are rho-independent

Question 12

What is a consensus sequence?

Answer 12

Sequence found most frequently in a survey of a large number of sequences

For promoters, the more similar a sequence is tot he consensus, the stronger the promoter

Question 13

What antibiotic is used to inhibit initiation of bacterial transcription?

Answer 13

Rifampin (rifamycin) - binds to B subunit of RNAP and inhibits the formation of the first phosphodiester bond

Question 14

What antibiotic is used to inhibit the elongation step of bacterial transcription?

Answer 14

Streptolydigin - binds to B subunit and blocks elongation

Question 15

What are the prokaryotic ribosomal subunits and their functions?

Answer 15

30S subunit - plays a role in translation initiation and has site for tRNA binding

50S subunit - peptide bond formation (transpeptidase)

Question 16

What are the four components of the translational machinery in bacteria?

Answer 16

Ribosomes

Soluble protein factors: Initiation factors, elongation factors, and release factors

tRNA

tRNA synthetases - catalyze the addition of AA to tRNA

Question 17

What are some features of the triplet code?

Answer 17

Universal for all organisms (with some minor exceptions)

Degenerate - more than one codon can code for a given AA

Three stop or nonsense codons (not recognized by any tRNA)

All translation begins at AUG which encodes methionine (N-formylmethionine)

Question 18

What are the stages of translation?

Answer 18

Initiation

Elongation

Termination

Question 19

What is the Shine-Dalgarno (S-D) sequence?

Answer 19

Also called the ribosome binding site

Sequence present in the 5’ nontranslated region of all prokaryotic mRNAs, has homology to the 3’ end of 16s rRNA

Question 20

What are the steps for translation elongation?

Answer 20

1: Charged tRNA enters A site
2: transpeptidation - transfer of peptidyl residue to free amino group of charged tRNA molecule in the A site
3: Deacylated tRNA leaves P site, peptidyl tRNA moves to P site

Question 21

How is translation terminated?

Answer 21

Soluble release factors recognize a stop codon, causing 70s to dissociate into 50S+30S and fall off mRNA

Question 22

What antibiotics block translation and how do they do so?

Answer 22

Streptomycin - blocks assembly of 70S initiation complex

Tetracyclines - block binding of charged tRNA to A site

Chloramphenicol - blocks peptidyltransferase reaction

Erythromycin - blocks translocation step

Question 23

What are three differences of transcription/translocation in prokaryotes?

Answer 23

Coupling can only occur in prokaryotes, eukaryotic nucleus separates the processes

Shine-Dalgarno sequence found only in prokaryotes

Many polycistronic transcripts for prokaryotes, very few in eukaryotes