R3 gene expression in prokaryotes

Question 1

What is the central dogma ?

Answer 1

The flow of information from DNA to RNA to proteins, but never backwards (formed by Francis Crick)

Question 2

What does coupled transcription and translation mean for prokaryotes ?

Answer 2

At one end, the mRNA is being synthesised whilst at the other it is being translated into proteins

Question 3

What does monocistronic and polycistronic mean ?

Answer 3

mono - one coding sequence for one protein, poly - multiple coding sequences for multiple proteins like an operon.

Question 4

What is the RBS ?

Answer 4

Ribosome binding site for the 30s subunit onto the mRNA strand.

Question 5

What is a STOP codon ?

Answer 5

they don’t code for an amino acid and direct the termination of the protein. UAA, UAG, UGA

Question 6

What is a holoenzyme ? and how does it form

Answer 6

The polymerase binds to a sigma factor creating a functional enzyme (the holoenzyme)

Question 7

what is a sigma factor ?

Answer 7

recognises the promoter site, then will release when primers start attaching.

Question 8

What do promoter sites usually end with ?

Answer 8

TATA box (TATATT)

Question 9

Describe in terms of equilibrium the relationship between RNA polymerase and unoccupied promoter

Answer 9

unoccupied promoter and free RNA polymerase are in an equilibrium to form the occupied promoter. The position of equilibrium is based on the affinity for the promoter.

Question 10

How can we detect the promoter strength ?

Answer 10

Reporter genes - encode for fluorescent proteins (GFP), stronger promoter = more GFP, brighter green colour

reporter gene codes for an enzyme - more transcription (lacZ), more beta-galactosidase - more product

Question 11

How does gene expression affect the final protein on each level ?

Answer 11

transcriptional level: more RNA transcripts –> more proteins

post transcriptional level: most stable RNA transcript –> more proteins

Translational level: premature translation termination –> less protein

post-translational level: protein degradation, activation or inactivation via phosphorylation, acetyle, sumo, ubquitin and other groups added

Question 12

What do the different boxes mean in the promoter ?

Answer 12

A different sigma factor binds to different sections of the the promoter (boxes)

sigma factors are an example of chemotaxis (response to a chemical)

Question 13

What is a regulon ?

Answer 13

A group of genes regulated as one unit, by activation or inhibition

Question 14

What is negative regulation?

Answer 14

The repressor binds to the operator region blocking the RNA polymerase from binding and transcribing downstream

Question 15

What is positive regulation?

Answer 15

Activator binds to the activator binding site further upstream from the promoter, anchoring the RNA polymerase and stopping it from leaving the promoter site. CAP protein

Question 16

What does beta-galactosidase do ?

Answer 16

Turns lactose into glucose and galactose

Question 17

Describe the regulation when lactose is present in the lac operon ?

Answer 17

The lactose is converted to alloLactose which allosterically inhibits the LacI protein, this detaches it from the operator allowing for RNA polymerase to bind and transcribe the operon. Negative regulation

Question 18

Describe the regulation when only glucose is present ?

Answer 18

The LacI protein blocks RNA polymerase from transcribing the operon

Question 18

Describe the regulation when only lactose is present and glucose is low?

Answer 18

Glucose usually inhibits ATP to cAMP but at low levels cAMP accumlates and binds to the CAP protein (Catabolite activator protein). This then can bind the the activator binding site, anchoring RNA polymerase to the promoter. Positive regulation

alloLactose inhibits LacI preventing it from blocking the promoter. negative regulation

Question 19

What is an example of allosteric activation and inhibition?

Answer 19

Activation - cAMP binds to the CAMP increasing its affinity to DNA

Inhibition - alloLactose binds to LacI, decreasing its affinity to DNA.

Question 20

What are the 4 uses for bacterial plasmid ?

Answer 20

Fertility, resistance to different chemicals, degradation of rare substances, virulence (ability to damage its host)

Question 21

What is conjugation in terms of prokaryote horizontal gene transfer ?

Answer 21

fertility plasmid transfer is coupled to DNA replication, the replicated plasmid is transferred to the other cell - this allows new traits like antibiotic resistance

Question 22

What is partial diploid in terms of prokaryote horizontal gene transfer ?

Answer 22

fertility plasmid integrates into the chromosome and then excise back, during the excision the adjacent chromosomal regions sometimes move to the plasmid

they excise due to the recipient cell already having genes present on the plasmid.