IGEM:MIT/2005/InsulinSensor

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Members

  1. Ray
  2. Jessica

Goals

  1. Detect CAP/CAMP
  2. Determine CAP/CAMP transfer function vs. glucose
  3. Universalize cAMP/CAP production given any input

Resources

Papers

  • Review:
    • Several hundred genes identified in E. coli regulated by cAMP/Crp complex (including lac and mal operons).
    • cAMP synthesis by adenylyl cyclase (AC) is the major regulatory site.
    • Detection of glucose levels by AC is incompletely understood but is approx. modelled as follows: in absence of glucose, members of the PTS system are normally phosphorylated, thus are continuously phosphorylating AC and driving cAMP production; in presence of glucose, members of the PTS system give up their phosphates to glucose and therefore primarily exist in nonphosphorylated forms, thus not stimulating AC and slowing cAMP production
    • Glucose transport DOES NOT correlate directly with AC activation. At medium levels of glucose, cAMP was still high despite PTS system working at saturation.
    • At low levels of glucose there is a second transition affecting cAMP levels, but it is different from strain to strain.
    • At low glucose levels, cAMP builds up inside to high levels and therefore has to be excreted (thus the transfer function of the extracellular cAMP closely resembles that of intracellular cAMP).
    • cAMP levels rise sharply when glucose levels drop to 0.3mM in minimal media (when PTS is working at 97% efficiency).
  • CcpA-Dependent Carbon Catabolite Repression in Bacteria
  • Review:
    • Carbon catabolite repression (CCR) follows different mechanisms in gram-postive and gram-negative bacteria
    • Gram positive uses CcpA dependent CCR
    • Gram negative uses cAMP-CRP
    • Gram postive modulates Ccpa with HPr as signalling itermediate
    • Gram negative uses EIIA-glc to activate adenyl cyclase and make cAMP. This is dependent on the phosphorylation state of EIIA-glc
    • Found homologies of CcpA signalling many proteobacteria and other types of bacteria

Books

Questions/Suggestions/Input from other team members

Thought this might be of interest... -KBS

Diagrams

The PTS and components with which it interacts --JTN

http://www.horizonpress.com/hsp/supplementary/pts/fig1.gif