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Substrate-induced conformational changes in the essential peripheral membrane-associated mannosyltransferase PimA from mycobacteria: implications for catalysis.

Abstract : Phosphatidyl-myo-inositol mannosyltransferase A (PimA) is an essential glycosyltransferase (GT) involved in the biosynthesis of phosphatidyl-myo-inositol mannosides (PIMs), which are key components of the mycobacterial cell envelope. PimA is the paradigm of a large family of peripheral membrane-binding GTs for which the molecular mechanism of substrate/membrane recognition and catalysis is still unknown. Strong evidence is provided showing that PimA undergoes significant conformational changes upon substrate binding. Specifically, the binding of the donor GDP-Man triggered an important interdomain rearrangement that stabilized the enzyme and generated the binding site for the acceptor substrate, phosphatidyl-myo-inositol (PI). The interaction of PimA with the beta-phosphate of GDP-Man was essential for this conformational change to occur. In contrast, binding of PI had the opposite effect, inducing the formation of a more relaxed complex with PimA. Interestingly, GDP-Man stabilized and PI destabilized PimA by a similar enthalpic amount, suggesting that they formed or disrupted an equivalent number of interactions within the PimA complexes. Furthermore, molecular docking and site-directed mutagenesis experiments provided novel insights into the architecture of the myo-inositol 1-phosphate binding site and the involvement of an essential amphiphatic alpha-helix in membrane binding. Altogether, our experimental data support a model wherein the flexibility and conformational transitions confer the adaptability of PimA to the donor and acceptor substrates, which seems to be of importance during catalysis. The proposed mechanism has implications for the comprehension of the peripheral membrane-binding GTs at the molecular level.
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Submitted on : Friday, August 27, 2010 - 11:37:41 AM
Last modification on : Wednesday, October 14, 2020 - 4:01:43 AM

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Marcelo E Guerin, Francis Schaeffer, Alain Chaffotte, Petra Gest, David Giganti, et al.. Substrate-induced conformational changes in the essential peripheral membrane-associated mannosyltransferase PimA from mycobacteria: implications for catalysis.. Journal of Biological Chemistry, American Society for Biochemistry and Molecular Biology, 2009, 284 (32), pp.21613-25. ⟨10.1074/jbc.M109.003947⟩. ⟨pasteur-00512066⟩

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