12. Regulatory Mechanisms II

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12. Regulatory Mechanisms II
2011-10-05 03:20:07
PMB 112 midterm1

general microbiology midterm 1
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  1. catabolite repression
    • allows bacterium to use the most efficient carbon source first
    • can be used in combination with specific regulatory mechanisms
  2. diauxic growth curve
    • cells grown on a mixed carbon source
    • lag in growth occurs when glucose is gone and cells must make new enzymes to use another carbon source
  3. relationship between glucose and cAMP with transcription
    • in the absence of glucose, transcription of genes for alternate sugar utilization is activated by CAP
    • CAP is activator, cAMP is co-activator
    • decrease in glucose -> increase in cAMP

    glucose present, [cAMP] low, entry of other sugars inhibited, CAP is off DNA, Lacl is on -> tx is repressed

    glucose absent, [cAMP] high, entry of other sugars permitted, CAP is on DNA, Lacl is off, tx is activated
  4. mechanism of glucose uptake
    • channel EnzIIc
    • phosphate passed PEP>EnzI>HPr>EnzIIA>EnzIIB>glucose
    • energy from bond of PEP
  5. PTS system when glucose is present
    • rapid entry of glucose
    • rapid flux of glycolytic pathway so PEP/pyruvate ratio is low

    • low phophorylation of PTS proteins
    • Enz IIa inhibits uptake of alternative sugars by binding to their transporters
  6. PTS system when glucose is absent
    • slow entry of glucose
    • slow flux through glycolytic pathway so PEP/pyruvate ratio is high

    • high phosphorylation of PTS proteins
    • Enz IIa~P binds and stimulates adenylate cyclase for the production of cAMP
  7. alternative sigma factors
    • E. coli has 7 different sigma factors:
    • σ70 - for most genes, normal growth, "housekeeping"
    • σ54 - nitrogen assimilation
    • σ38 (σs) - stationary phase, various stress conditions
    • σ32 (σH) - heat shock response
    • σ28 - flagellum biosynthesis
    • σ24 (σE) - response to misfolded proteins in perplasm, cell envelope trouble
    • σ19 - for iron transport genes

    the cell regulates global gene expression patterns by controlling which sigma factors are present and active
  8. how do cells respond to and recover from heat shock?
    • protein denaturation
    • cells synthesize chaperones and proteases to refold or degrade damaged proteins
    • heat shock-induced genes have promoter sequences that bind to σ32 (rpoH)
  9. heat shock genes under normal conditions
  10. heat shock conditions
    • increases the amount and activity of σ32
    • stem-loop melted
    • translation is efficient because mRNA secondary structure has been removed
    • DnaK chaperone binds heat-denatured proteins and promotes their refolding or degradation
    • σ32 is not inactivated or degraded by binding to DnaK
    • interacts with RNAP core to promote tx of heat shock-induced genes
  11. recovery from heat shock
    • one of the genes turned on by σ32 is dnaK itself, creating a homeostatic mechanism that returns the cell to its normal state
    • over time, denatured proteins are refolding, DnaK levels rise, and there is enough DnaK to rebind σ32
    • refolded proteins released
    • σ32 inactivated and/or degraded by FtsH
  12. SOS response to DNA damage
    • RecA binds to ssDNA exposed by UV damage and stalled replication forks
    • RecA+ssDNA stimulates LexA repressor auto-cleavage, LexA-repressed genes are turned on
    • SulA protein inhibits cell division
    • UmuCD now induced, can perform mutagenic translesion DNA synthesis by using damaged DNA strand as template and adds random bases
  13. How do cells reset after the DNA damage is repaired?
    • in response to stress or damage, cells activate tx of new genes
    • after the stress/damage is over, cells reset by degrading or deactivating the proteins that respond to the stress, because they often aren't beneficial in normal conditions

    SulA protein is degraded by the Lon protease - sulA gene is repressed by new LexA, cell division will restart

    UmuCD is degraded by ClpXp protease - UmuCD tx stops, no more mutagenic replication occurs
  14. two-component signal transduction
    • sensor kinase protein:
    • histidine kinases in cytoplasmic membrane autophosphorylate or external signal dimerization cross-phosphorylation
    • phosphoryl group is transferred to an Asp residue on a response regulator which activates the protein

    • response regulator proteins can:
    • activates or represses tx
    • output domains can be enzymes turned on/off by phosphorylation of receiver domain
    • sometimes a receiver domain acts alone by binding to a downstream protein
  15. examples of two-component signaling pathways that are global transcriptional regulators
    • ArcB kinase-ArcA response regulator:
    • ArcB kinase sense oxygen levels and autophophorylates in low oxygen
    • ArcB phophorylates ArcA, which turns off/on a large set of genes

    • NtrB kinase-NtrC response regulator :
    • NtrB sense nitrogen levels and phosphorylates in response to nitrogen starvation
    • NrtB phosphorylates NtrC which turns off/on genes