Rewiring the specificity of two component signal transduction system
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Rewiring the Specificity of Two-Component Signal Transduction System. By Jeffrey M. Skerker , Barrett S. Perchuk , Albert SiryaPorn , Emma A. Lubin , Orr Ashenberg , Mark Goulian , and Michael T. Laub Cell (2008); 133(6); 1043-1054 Presented by Jose R. Echevarria for 20.385 02-24-2010.

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Rewiring the specificity of two component signal transduction system

Rewiring the Specificity of Two-Component Signal Transduction System

By Jeffrey M. Skerker, Barrett S. Perchuk, Albert SiryaPorn, Emma A. Lubin, Orr Ashenberg, Mark Goulian, and Michael T. Laub

Cell (2008); 133(6); 1043-1054

Presented by Jose R. Echevarria for 20.385

02-24-2010


Hypothesis
Hypothesis Transduction System

Could the Specificity of HistidineKinases be determined by amino acids that exhibit a large degree of covariation.

If you vary a subset of amino acids of HistidineKinases, could that modify the specificity of the HistidineKinases.


Histidine kinase diagram
Histidine Transduction SystemKinase Diagram


Structural analysis based on histidine phosphotransferase
Structural Analysis based on Transduction SystemHistidinePhosphotransferase

Residues do not perfectly correlate with contact residues of the Spo0B:Spo0F model


Primary alignments
Primary Alignments Transduction System


Search for covarying residues
Search for covarying residues Transduction System


Search for covarying residues1
Search for Covarying Residues Transduction System

  • 43 amino acids found with an MI greater than .35

  • 36 of them included the dHp domain of histidinekinase and it’s response regulator

  • Formula for MI:


Dhp domain is critical
DHp Transduction System Domain is Critical


7 residue chimera construct
7 Residue Chimera Construct Transduction System

Mutants with varying levels of homology to the EnvZDHp domain.

Holds 7 Specificity – determining residues at the C- terminal, N-terminal portion of helix 2, and a connecting loop.


More chimera testing
More Chimera Testing Transduction System


K cat k m analysis
K Transduction Systemcat/Km Analysis



New mutants containing loop
New Mutants containing Loop Transduction System


Well behaved chimeras
Well behaved Chimeras Transduction System


In vivo testing
In Vivo Testing Transduction System


Critique
Critique Transduction System

Key Assumptions

  • Phylogeny is greatly reduced by screening for genomes with less than 90 % identity.

  • Covariance determines the amino acids that would be more deterministic in histidine specificity.

    Pit-Fall

  • Need a combination of approaches for better understanding.


Significance
Significance Transduction System

  • Proof of the possibility to rewire the specificity of input systems by modifying a small number of residue.

  • Model through which we can add complexity and apply to other biological systems in vivo.

  • Cross-talk Suppression between the CpxA-CpxR and EnvZ-OmpR Two-Component Systems in E. coli; Albert Siryaporn & Mark Goulian


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