Regulation of transcription/translation in prokaryotes

Question 1

what is rRNA?

Answer 1

ribosomal RNA

Question 2

what is the function of rRNA?

Answer 2

forms part of the ribosomes structure (integral part of the architecture)

Question 3

what is the function of tRNA?

Answer 3

delivers amino acids to the ribosome during protein synthesis - also known as adaptor molecules

Question 4

what are the three types of functional RNA in a bacterial cell?

Answer 4

• mRNA
• tRNA
• rRNA

Question 5

how are the RNA in bacterial cells synthesised?

Answer 5

synthesised by DNA dependent RNA polymerase (i.e. require a gene that codes for that RNA)

Question 6

what are the 5 subunits of the RNA polymerase core enzyme?

Answer 6

alpha
alpha
beta
beta prime (')
omega (w)

Question 7

what is the holoenzyme?

Answer 7

core enzyme plus sigma subunit/factor

Question 8

what is the function of the beta subunits?

Answer 8

interact/make contact with non-specific DNA and RNA (sigma is the one that confers specificity by promoter recognition)
Beta prime is involved in DNA binding and beta is involved in initiation and chain elongation

Question 9

what is an ORF?

Answer 9

open reading frame - portion of DNA that contains no stop codons when transcribed
promoters are ORFs

Question 10

what are the two things that happen when the sigma factor binds to the core enzyme?

Answer 10

1. RNA polymerases affinity for non-specific (i.e. non-promoter) DNA is reduced
2. Its affinity for promoters (i.e. specific DNA) is simultaneously increased

Question 11

how does the holoenzyme find the promoter sequence?

Answer 11

via diffusion search , also known as scanning. The holoenzyme has an unusual crab claw structure that allows it to move up and down the DNA sequence
.

Question 12

what is the proposed mechanism of RNA pol diffusion search?

Answer 12

the structure of the polymerase is like a hand. The core enzyme has a closed conformation while the holoenzyme (i.e. when sigma binds) has an open conformation allowing it to move across DNA and scan for DNA
- binds to non-target DNA and finds promoter by sliding search

Question 13

what are the 5 ways RNA polymerase can scan double-stranded DNA?

Answer 13

1. 1D diffusion/translocation (not moving from DNA)
2. hopping along the strand
3. jumping
4. 3D diffusion (leaving DNA and reattaching elsewhere
5. intersegment transfer - jumping from one loop of DNA to another loop of close proximity (which may actually be many sequences up/downstream - folding of DNA brings them closer)

Question 14

what is a consensus sequence?

Answer 14

a theoretical sequence consisting of the most frequently occurring codons /amino acids at each site. Deduced by aligning sequences (e.g. different promoters) and looking at the occurrence of bases at a certain site. The closer in sequence a promoter is to the consensus the greater the level of transcription.

Question 15

give examples of consensus sequences in e.coli

Answer 15

-10 and -35

Question 16

what is +1?

Answer 16

the start of transcription

Question 17

how is transcription directionality conferred?

Answer 17

-35 and -10 boxes are arranged asymmetrically within the promoter to position the polymerase in the correct orientation.

Question 18

how does the sigma factor recognise -10 and -35 regions?

Answer 18

domain 2 of sigma binds to -10 which helps separate the DNA strands (promoter melting)

• 10 has an extended region of two bases that is recognised by domain 3
• domain 4 recognises -35 and is attached to the flexible part of the core enzyme so it can accommodate different lengths of space between the -35 and -10 regions

Question 19

what are early genes?

Answer 19

genes that are transcribed before the replication of phage DNA. they code for regulators and proteins needed at later stages of infection.

Question 20

what are middle genes?

Answer 20

phage genes that are regulated by the proteins coded by early genes. e.g. proteins that catalyse replication of phage DNA or regulated expression of later genes

Question 21

what are late genes?

Answer 21

genes transcribed when phage DNA is being replicated. they code for components of the phage particle