Ra. Buono et al., INFRARED SPECTROSCOPIC STUDIES OF CALCIUM-BINDING TO INHIBITED BETA-TRYPSINS, Biochimica et biophysica acta. Protein structure and molecular enzymology, 1204(1), 1994, pp. 124-128
Fourier transform infrared spectroscopy was used to examine the effect
of calcium binding on the secondary structure of two inhibited bovine
beta-trypsins. Neither the diisopropyl fluorophosphate- nor benzamidi
ne-inhibited forms showed detectable secondary structure perturbation
upon calcium binding at pD 6.9 and 5.0, respectively. Considered in li
ght of the recent assignment of an amide I' band to the autolysis loop
of bovine beta-trypsin, these results contradict the generally held h
ypothesis that calcium slows trypsin autolysis by induction of a confo
rmational change at this site and support the recent contention that t
he mechanism of action has a specific electrostatic origin. In additio
n, the appearance of a band at 1699 cm(-1) in the benzamidine-inhibite
d form can be interpreted as resulting from the NC-N stretching vibrat
ions of the amidinium moiety, which the observed crystal structure ind
icates is hydrogen-bonded to the carboxyl group of active-site Asp-189
.