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  current news   Press   selected story    
     
  21th October 2010  
 

Structural Basis of the Sensor-Synthase Interaction in Autoinduction of the Quorum Sensing Signal DSF Biosynthesis

 
 




Authors:
Zhihong Cheng1,5, Ya-Wen He1,5, Siew Choo Lim1,4,5, Rohini Qamra1,5, Martin A. Walsh2 Lian-Hui Zhang3 and Haiwei Song1,3,4.

1 - Institute of Molecular and Cell Biology, 61 Biopolis Drive, Proteos, Singapore 138673.
2 - 2MRC France, CRG BM14, ESRF, B.P.220, F-38043, Grenoble CEDEX, France.
3 - Department of Biological Sciences, National University of Singapore, 14 Science Drive, Singapore 117543.
4 - School of Biological Sciences, Nanyang Technological University, 60 Nanyang Drive, Singapore 637551.
5 - These authors contributed equally.


Published in Structure, 2010 Sept 8, 18(9): 199-209.

Abstract
The diffusible signal factor (DSF)-dependent quorum sensing (QS) system adopts a novel protein-protein interaction mechanism to auto-regulate the production of signal DSF. Here we present the crystal structures of DSF synthase RpfF and its complex with the REC domain of sensor protein RpfC. RpfF is structurally similarity to the members of the crotonase superfamily and contains an N-terminal α/β spiral core domain and a C-terminal α-helical region. Further structural and mutational analysis identified two catalytic glutamate residues, which is the conserved feature of the enoyl-CoA hydratases/dehydratases. A putative substrate-binding pocket was unveiled and the key roles of the residues implicated in substrate binding were verified by mutational analysis. The binding of the REC domain may lock RpfF in an inactive conformation by blocking the entrance of substrate binding pocket, thereby negatively regulating DSF production. These findings provide a structural model for the RpfC-RpfF interaction-mediated novel QS autoinduction mechanism.

 
 


 
 


Figure legend: Structure of RpfF alone and in complex with the REC domain of RpfC.
Ribbon diagrams showing the self-association fold of RpfF (A) and the overall structure of the RpfF/REC complex (B). (C) Mutagenesis reveals the critical residues involved in catalytic activity. Upper panel: TLC plate was used to quantify DSF activity as indicated by the presence of a blue zone. Middle panel: the amount of DSF production in WT and mutant RpfF. Lower panel: Western blot analysis to check expression of WT and mutant RpfF. (D) Cavity analysis of RpfF showed that the cavity in the putative catalytic site is too small to accommodate a substrate with the same carbon chain length as DSF (12 carbon atoms). The stick model in orange color is octanoyl-CoA from PDB code: 2DUB. The stick model in slate color is pre-DSF-CoA, which is modeled using the structure of octanoyl-CoA as a template, and is obviously larger than the cavity, suggesting that a conformation change will occur when RpfF binds to substrate.

For more information on Lianhui ZHANG’s lab, please click here.

For more information on Haiwei SONG’s lab, please click here.