The Active Site Protonation States of the class A CTX-M-type perdeuterated Toho-1 -lactamase Determined by Neutron Diffraction Support a Role for Glu166 as the General Base in Acylation

• Published in: FEBS letters Vol. 1; no. 1
• Main Authors: Coates, Leighton, Cooper, Jon, Tomanicek, Stephen J, Blakeley, Matthew P., Chen, Yu, Wang, Kathy K
• Format: Journal Article
• Language: English
• Published: United States 01-01-2010
• Subjects: ACYLATION; ATOMS; DEUTERIUM; MUTANTS; NEUTRON DIFFRACTION; NEUTRONS; PHYSICS OF ELEMENTARY PARTICLES AND FIELDS
• ISSN: 0014-5793; 1873-3468

Room temperature neutron diffraction data of the fully perdeuterated Toho-1 R274N/R276N double mutant {beta}-lactamase in the apo form were used to visualize deuterium atoms within the active site of the enzyme. This perdeuterated neutron structure of the Toho-1 R274N/R276N reveals the clearest picture yet of the ground-state active site protonation states and the complete hydrogen-bonding network in a {beta}-lactamase enzyme. The ground-state active site protonation states detailed in this neutron diffraction study are consistent with previous high-resolution X-ray studies that support the role of Glu166 as the general base during the acylation reaction in the class A {beta}-lactamase reaction pathway.