HYDROGEN-DEUTERIUM EXCHANGE OF TRYPSIN AND TRYPSIN DERIVATIVES

Hydrogen-deuterium exchange of trypsin and two trypsin derivatives, diisopropylphosphoryltrypsin and N-tosyl-L-lysinè chloromethyl ketone-trypsin, was studied by means of infrared spectroscopy. Measurements were made at 23° (0.1 M KC1 in D2O) and at pD values from 2 to 8. Four different classes of exchanging hydrogens were found: a fast class with a pD-dependent rate constant for exchange, a medium class with an essentially pD-independent rate constant for exchange, a slow class with a pD-independent rate constant for exchange, and a “core” class which does not exchange within 24 hr. The fast class could only be observed at pD values around 2 and probably consists of completely exposed peptide hydrogens. The medium and slow classes appear to exchange by a mechanism in which the rate is determined by transconformational exposure of unexposed peptide groups. The rate constants for exchange of the medium or slow classes are essentially the same for trypsin, diisopropylphosphoryltrypsin, and N-tosyl-L-lysine chloromethyl ketone-trypsin. The numbers of hydrogens per class are essentially the same for the medium, slow or “core” classes for all three substances. These results imply that the medium slow, or “core” peptide hydrogens are essentially structurally identical. © 1969, American Chemical Society. All rights reserved.