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Molecular Basis of Membrane Association by the Phosphatidylinositol Mannosyltransferase PimA Enzyme from Mycobacteria

Abstract : Phosphatidyl-myo-inositol mannosyltransferase A (PimA) is an essential glycosyltransferase that initiates the biosynthetic pathway of phosphatidyl-myo-inositol mannoside, lipomannan, and lipoarabinomannan, which are key glycolipids/lipoglycans of the mycobacterial cell envelope. PimA belongs to a large family of membrane-associated glycosyltransferases for which the understanding of the molecular mechanism and conformational changes that govern substrate/membrane recognition and catalysis remains a major challenge. Here, we determined that PimA preferentially binds to negatively charged phosphatidyl-myo-inositol substrate and non-substrate membrane model systems (small unilamellar vesicle) through its N-terminal domain, inducing an important structural reorganization of anionic phospholipids. By using a combination of single-point mutagen-esis, circular dichroism, and a variety of fluorescence spectros-copy techniques, we determined that this interaction is mainly mediated by an amphipathic-helix (2), which undergoes a substantial conformational change and localizes in the vicinity of the negatively charged lipid headgroups and the very first carbon atoms of the acyl chains, at the PimA-phospholipid interface. Interestingly, a flexible region within the N-terminal domain, which undergoes-strand-to-helix and-helix-to-strand transitions during catalysis, interacts with anionic phospholipids; however, the effect is markedly less pronounced to that observed for the amphipathic 2, likely reflecting structural plasticity/variability. Altogether, we propose a model in which conformational transitions observed in PimA might reflect a molten globule state that confers to PimA, a higher affinity toward the dynamic and highly fluctuating lipid bilayer.
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Submitted on : Wednesday, October 5, 2016 - 4:05:09 PM
Last modification on : Thursday, June 11, 2020 - 12:00:05 PM

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Ane Rodrigo-Unzueta, Mariano A. Martínez, Natalia Comino, Pedro M. Alzari, Alexandre Chenal, et al.. Molecular Basis of Membrane Association by the Phosphatidylinositol Mannosyltransferase PimA Enzyme from Mycobacteria. Journal of Biological Chemistry, American Society for Biochemistry and Molecular Biology, 2016, 291 (27), pp.13955 - 13963. ⟨10.1074/jbc.M116.723676⟩. ⟨pasteur-01376787⟩

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