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New insight into the catalytic mechanism of bacterial MraY from enzyme kinetics and docking studies

Liu, Yao and Rodrigues, Joao P. G. L. M. and Bonvin, Alexandre M. J. J. and Zaal, Esther A. and Berkers, Celia R. and Heger, Michal and Gawarecka, Katarzyna and Swiezewska, Ewa and Breukink, Eefjan and Egmond, Maarten R. (2016) New insight into the catalytic mechanism of bacterial MraY from enzyme kinetics and docking studies. Journal of Biological Chemistry, 291 (29). pp. 15057-15068. ISSN 0021-9258


Official URL: http://www.jbc.org/content/291/29/15057.long


Phospho-MurNAc-pentapeptide translocase (MraY) catalyzes the synthesis of Lipid I, a bacterial peptidoglycan precursor. As such, MraY is essential for bacterial survival and therefore is an ideal target for developing novel antibiotics. However, the understanding of its catalytic mechanism, despite the recently determined crystal structure, remains limited. In the present study, the kinetic properties of Bacillus subtilis MraY (BsMraY) were investigated by fluorescence enhancement using dansylated UDP-Mur-NAc-pentapeptide and heptaprenyl phosphate (C35-P, shortchain homolog of undecaprenyl phosphate, the endogenous substrate ofMraY)as second substrate. Varying the concentrations of both of these substrates and fitting the kinetics data to two-substrate models showed that the concomitant binding of both UDPMurNAc-pentapeptide-DNS and C35-P to the enzyme is required before the release of the two products, Lipid I andUMP.We built a model of BsMraY and performed docking studies with the substrate C35-P to further deepen our understanding of how MraY accommodates this lipid substrate. Based on these modeling studies, a novel catalytic role was put forward for a fully conserved histidine residue inMraY(His-289 in BsMraY), which has been experimentally confirmed to be essential for MraY activity. Using the current model of BsMraY, we propose that a small conformational change is necessary to relocate the His-289 residue, such that the translocase reaction can proceed via a nucleophilic attack of the phosphate moiety of C35-P on bound UDP-MurNAc-pentapeptide.

Item Type:Article
Subjects:Q Science > QH Natural history > QH301 Biology
Divisions:Department of Lipid Biochemistry
ID Code:1241
Deposited By: Ewa Swiezewska
Deposited On:15 Nov 2016 12:00
Last Modified:08 Mar 2018 15:34

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