Science, Vol 259, Issue 5095, 669-673
Copyright © 1993 by American Association for the Advancement of Science
The hydrolytic water molecule in trypsin, revealed by time-resolved Laue crystallography
PT Singer,
A Smalas,
RP Carty,
WF Mangel,
and
RM Sweet
Biology Department, Argonne National Laboratory, IL 60439.
Crystals of bovine trypsin were acylated at the reactive residue, serine 195, to form the transiently stable p-guanidinobenzoate. Hydrolysis of this species was triggered in the crystals by a jump in pH. The hydrolysis was monitored by three-dimensional Laue crystallography, resulting in three x-ray diffraction structures, all from the same crystal and each representing approximately 5 seconds of x-ray exposure. The structures were analyzed at a nominal resolution of 1.8 angstroms and were of sufficient quality to reproduce subtle features in the electron-density maps for each of the structures. Comparison of the structures before and after the pH jump reveals that a water molecule has positioned itself to attack the acyl group in the initial step of the hydrolysis of this transient intermediate.
