The hydrolytic water molecule in trypsin, revealed by time-resolved Laue crystallography.
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.
Singer
P T
PT
Biology Department, Argonne National Laboratory, IL 60439.
Smalås
A
A
Carty
R P
RP
Mangel
W F
WF
Sweet
R M
RM
D000595:Amino Acid Sequence; D000818:Animals; D002417:Cattle; D003461:Crystallography / Q000379:methods; D007202:Indicators and Reagents; D008958:Models, Molecular; D011487:Protein Conformation; D012694:Serine; D014357:Trypsin / Q000737:chemistry*; D014867:Water
D007202:Indicators and Reagents; D014867:Water; D012694:Serine; D014357:Trypsin