Protonation state and substrate binding to B2 metallo-b-lactamase CphA from Aeromonas hydrofila.

F. SIMONA, A. MAGISTRATO, D. M. A. VERA, G. GARAU, A. J. VILA, P. CARLONI.

Proteins: Structure, Function, and Bioinformatics

Wiley Interscience

Lugar: New York; Año: 2007 vol. 69 p. 595 - 695

0887-3585

<!-- @page { size: 21.59cm 27.94cm; margin: 2cm } P { margin-bottom: 0.21cm } --> The zinc enzymes metallo b-lactamases coun- teract the beneficial action of b-lactam anti- biotics against bacterial infections, by hydro- lyzing their b-lactam rings. To understand structure/function relationships on a repre- sentative member of this class, the B2 MbL CphA, we have investigated the H-bond pat- tern at the Zn enzymatic active site and sub- strate binding mode by molecular simulation methods. Extensive QM calculations at the DFT-BLYP level on eleven models of the pro- tein active site, along with MD simulations of the protein in aqueous solution, allow us to propose two plausible protonation states for the unbound enzyme, which are probably in equilibrium. Docking procedures along with MD simulations and QM calculations suggest that in the complex between the enzyme and its substrate (biapenem), the lat- ter is stable in only one of the two protona- tion states, in addition it exhibits two differ- ent binding modes, of which only one agrees with previous proposals. In both cases, the substrate is polarized as in aqueous solution. We conclude that addressing mechanistic issues on this class of enzymes requires a careful procedure to assign protonation states and substrate docking modes.