Transcript Regulation of Gene expression
Regulation of Gene expression by E. Börje Lindström
This learning object has been funded by the European Commissions FP6 BioMinE project
Introduction • Biosynthetic reactions consume energy: Sophisticated control mechanisms in bacteria • Available energy is limited in Nature: Production of as much cell material per energy as possible • The environment is important: • Two types of model systems: - the nutrient in the medium is used first - rapid and drastic changes in the nutrients - reversible control reactions needed - Biosynthetic - Catabolic
Biosynthetic reactions Tryptophan is chosen as a model system: - Tryptophan is an essential amino acid - Tryptophan is missing in some plant proteins - of industrial importance • The bacterial cells are controlling the biosynthesis of tryptophan in three ways: - feedback
inhibition
- end product
repression
-
attenuation
Biosynthetic reactions, cont.
• Feedback inhibition: - The biosynthesis of tryptophan occurs in several steps: Chorismate + glutamine antranilic acid B C D tryptophan Mechanism: - enzyme E1 (the first enzyme) is an allosteric protein with - a
binding site
for for the
substrate
- a
binding site
for the
effectors
(inhibitor = try) • E1 + try [E1-try]-complex that is inactive • the complete biosynthesis of try is stopped
Biosynthetic reactions, cont.
• End product repression (EPR): - In spite of ’end product inhibition’ -
loss of energy
due to enzymes E2-E5 are still synthesized - another regulation is needed -
end product repression
Biosynthetic reactions, cont.
Mechanism:
P O att E1 E2 E3 E4 E5
P = promoter; O = operator att = attenuator • RNA polymerase binds to
P
E1 – E5
= structural genes for the enzymes E1-E5.
Initiation of mRNA synthesis • The repressor is an allosteric protein -
inactive
without tryptophan (does not bind to the operator) • tryptophan acts as co-repressor -binds to the repressor • The repressor binds to
O
- makes the
repressor active
Blocks the RNA polymerase movement
Biosynthetic reactions, cont.
• Attenuator region: - barrier for the RNA polymerase 1) + try the polymerase removed from the DNA 2) - try the polymerase continues into the structural genes • EPR inhibits all enzymes in tryptophan biosynthesis save energy - however, a slow total inhibition – does not effect already existing enzymes - high specificity – only the tryptophan operon is effected
Biosynthetic reactions, cont.
Biosynthetic reactions, cont.
Biosynthetic reactions, cont.
Biosynthetic reactions, cont.
Catabolic reactions • Catabolic systems are
inducible
• The inducer is the available carbon/energy source • Model system – lactose operon in
E. coli R P O lac
Z
lac
Y
lac
A • Where: - gene
R :
repressor protein –
active
without the inducer blocks mRNA polymerase - gene
lac
Z : b -galactosidase – splits lactose into glycose + galactose - gene
lac
Y: permease – transport lactose into the cell -
no attenuator
sequence in catabolic systems
Catabolic reactions, cont.
• Mechanism:
+ lactose:
- transported into the cell transformed into allo-lactose (inducer) - allo-lactose + repressor [allo-lactose-repressor]- complex
inactive
- RNA polymerase starts transcription of lactose operon b -galactosidase is produced break down of lactose
- lactose:
-[allo-lactose-repressor]- complex
disintegrate
- the repressor binds to
O
and blocks further transcription of the operon
Catabolic reactions, cont.
Catabolic reactions, cont.
Catabolic repression (glucose effect) • Works in bacteria and other prokaryotes (here in
E. Coli
K12) • Diauxi: - growth on two energy sources
glucose + lactose
- two-step growth curve Log OD Growth on lactose lactose Growth on glucose time glucose
Catabolic repression (glucose effect) • Mechanism: -cAMP an important substance - required for initiation of transcription of many inducible systems - global regulation - glucose present [cAMP] (decreases) - CAP (
k
atabolite
a
ctivator
p
rotein) an allosteric protein - [cAMP-CAP]-complex binds to the promoter promotes transcription -production of b -galactosidase -1) lactose present - 2) [cAMP-CAP]-complex present
Catabolic repression (glucose effect), cont.
• + glucose: -
no
[cAMP-CAP]-complex -
no
transcription of lactose operon -
no
b -galactosidase production • - glucose: - [cAMP-CAP]-complex present - transcription of lactose operon b -galactosidase production - brake down of lactose
Catabolic repression (glucose effect), cont.
• Conclusions: -
Katabolite repression
– a very useful function in bacteria - forces the bacteria to
use the best energy source
first
Other types of Regulations • Constitutive systems: - no regulation - always present - Enzymes that are needed during all types of growth - e.g. those involved in glycolysis • mRNA: - Unstable - half-life ~ 2 min sub-units new mRNA • polycistronic mRNA - one operator for several genes • monocistronic mRNA one operator per gene (in eukaryotes)