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Science 29 January 1993:
Vol. 259. no. 5095, pp. 669 - 673
DOI: 10.1126/science.8430314
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Articles

Science, Vol 259, Issue 5095, 669-673
Copyright © 1993 by American Association for the Advancement of Science

articles

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.


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Response.
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