Lecture 2

Question 1

Do bacteria need to constantly produce proteins?

Answer 1

Bacteria do not need to make all of their proteins all the time, their requirements depend on available food sources and other external conditions.

Question 2

What proteins do E.coli need to make?

Answer 2

They need to transcribe genes to make proteins to use the food substrates found nearby, in the gut it depends on the host’s food choices.
Genes encoding enzymes that make amino acids that are absent in the environment.

Question 3

What are the 2 main control mechanisms of transcription?

Answer 3

Choosing which genes to start transcribing

Deciding where to stop transcribing

Question 4

Control: choosing which genes to start transcribing

Answer 4

Alternative sigma factors or operons

Question 5

Control: deciding where to stop transcribing

Answer 5

Attenuation or anti-terminators

Question 6

Sigma factors (control)

Answer 6

70 sigma factors can be recognised. Sigma 70 switches on most housekeeping genes. The sigma factor recognises the -35 box. If a different sigma factor is used by the RNA polymerase it will recognise a different sequence. Alternative sigma factors are used to switch on/off different groups of genes in bacteria.

Question 7

What sigma factors recognise genes activated by heat shock?

Answer 7

These genes have a different -35 and -10 box sequence so are not recognised by sigma 70, instead it is recognised by sigma 32.

Question 8

What is heat shock and what events follow it?

Answer 8

Slightly above optimum temperature where proteins begin to denature and there is an accumulation of misfolded proteins. Proteases must be produced to break down misfolded proteins or chaperons are produced to correctly refold the misfolded protein.
When unfolded proteins are detected sigma 32 is produced so genes can be switched on.

Question 9

What happens when you add a different sigma factor to the core RNA polymerase enzyme.

Answer 9

A different version of the holoenzyme is made with different recognition capabilities.
Production of alternative sigma factors is induced by different conditions (heat shock)

Question 10

Describe what most sigma factors have in common

Answer 10

Most are part of the sigma 70 family. They all recognise -35 and -10 regions

Question 11

What is an operon (control)?

Answer 11

Collection of genes that produce proteins that are all needed at the same time as they are controlled by the same promoter- they can be switched off when they are not needed.

Question 12

Operons allow _____ control of expression in prokaryotes.

All ____ in an operon are under the _____ control.

Answer 12

Co-ordinate
Genes
Same

Question 13

Where is the operator region?

Answer 13

The operator region is in the promoter region.

Question 14

What will the operon transcribe?

Answer 14

A polycistronic mRNA molceule when switched on which is mRNA containing all the information for the genes.

Question 15

Describe the E.coli genome

Answer 15

Circular
4639 Kbp
Many genes of related function are grouped together

Question 16

What are the two groups of genes that are regulated together?

Answer 16

Catabolic- proteins encoded are involved in the breakdown of substrates e.g. the lac operon
Biosynthetic - proteins encoded are involved in building new molecules e.g. Trp operon

Question 17

Positive and negative mechanisms of control.

Answer 17

Positive operons will facilitate transcription and negative operons prevent transcription.

Question 18

Give an example of a catabolic operon and a biosynthetic operon

Answer 18

Catabolic: lac operon
Biosynthetic: Trp operon

Question 19

What is a biosynthetic operon?

Answer 19

Expression of enzymes that together synthesise small molecules (amino acids, nucleotides). If these small molecules are not in the growth media the bacterium has to make them so the operon is switched on.

Question 20

Describe the genes that make up the Trp operon

Answer 20

5 structural genes: trpA, trpB, trpC, trpD, trp E

These encode 5 enzymes that are in the biosynthetic pathway that leads to the production of a tryptophan operon.

Question 21

What codes the repressor protein in the Trp operon?

Answer 21

trpR

Question 22

Describe the repressors in the Trp operon

Answer 22

apo-repressor: Trp repressor

Holo-repressor: Trp repressor & tryptophan

Question 23

What happens when there are low levels of tryptophan?

Answer 23

Tryptophan galls off of the trp repressor and repression is stopped. Expression of operon occurs

Question 24

Positive control

Answer 24

Activator molecule binds to an operator region and transcription occurs.
Repression occurs when the activator molecule in inactivated & detaches from the gene

Question 25

Negative control

Answer 25

Repressor molecule binds to an operator region to stop transcription.
For transcription to occur the repressor needs to be inactivated and detach from the gene.

Question 26

Describe what happens in the Arabinose operon if arabinose is present.

Answer 26

In the presence of arabinose the operon needs to be switched on.
CAP +cAMP bind
AraC protein and arabinose bind. These act as an activator to promote transcription.

Question 27

Describe what happens in the Arabinose operon if arabinose is absent.

Answer 27

AraC protein will undergo a conformation change so it can bind the O & I region. The operon needs to loop to allow this binding and transcription is obstructed.

Question 28

What is significant about the arabinose operon?

Answer 28

Dual control - positive and negative control

Question 29

What are the genes involved in the arabinose operon?

Answer 29

AraC: control gene for activator protein
AraO & AraI are the control sires (initiator region)
BAD: structural genes which are involved in the conversion of sugar arabinose to D-xylase-5-phosphate
B => Kinase
A => Isomerase
D => Epimerase