Lec14_HTP Regulon (Prokaryote?)

Question 1

Heat shock

Answer 1

sudden increase of the microbes’ environment temp
increase synthesis of heat shock proteins

heat shock rxn

• conserved in all organisms
• provides organism with temporary resistance to damaging effects of heat
• other conditions also bring about heat shock - stress rxn

E. coli genome

• only two genes found in operons coding for heat shock proteins
• Dnaj and DnaK

Question 2

what happens to proteins during high temp and how does heat shock proteins protect cell

Answer 2

at high temp

• some proteins denature (lose 3D conformation)
• or become less function

Heat shock proteins protect cell by:

1. destroying denaturing proteins
2. acts as a chaperone -3D
3. accompany proteins to specific cellular locations, where they are needed

Question 3

Induction of HTP regulon - Sigma factor
AND
extent to which proteins are synthesized

Answer 3

activator - sigma factor 32

sigma factor 32

• binds to core enzyme of RNA Polymerase to form holoenzyme
• recognizes promoters of genes in HTP regulon
• recognize -35 and -10 DNA sequences that differs from that of sigma factor 70

extent to which proteins are synthesized
- depends on extent of temp increase that taken place

Question 4

Sigma factor 32

Answer 4

normally present in cell

• smaller quantities than sigma factor 70
• required for transcription of certain genes under normal circumstances
• encoded by rpoH gene
• Promoter 3 recognize by sigma factor 24 (only activate at high temp) -responsible for high induction of rpoH transcription

Question 5

rpoH gene

Answer 5

• rpoH null mutant = not able to grow at temp above 20’C
• complex initiation area
  *different promoters
  *P1,P4 and P5 - recognize by sigma factor 70
  *P3 - recognize by sigma factor 24
  sigma factor 24
• only active at high temp
  -responsible for high induction of rpoH transcription

Question 6

HTP regulon regulation

Answer 6

No stress
(absence of denatured proteins)
- heat shock proteins, DnaJ and DnaK associate with sigma factor 32
- causes sigma factor 32 to destabilize = short half life

Stress

• presence of denatured proteins
• denatured proteins associate with DnaJ and DnaK
• increases sigma factor 32 half-life
• increasing transcription of HTP regulon genes
• when denatured proteins disappear
• sigma factor 32 levels drop
• heat shock rxn returns to normal

Question 7

DnaJ and DnaK binding affinity

Answer 7

for denetured proteins are higher than for sigma factor 32