Hydrogen-deuterium exchange in the black-eyed pea trypsin and chymotrypsin inhibitor and its complex with beta-trypsin.
The H-D exchange of the black-eyed pea trypsin and chymotrypsin inhibitor (BTCI) in D2O was studied by an ultraviolet spectroscopic method recently proposed (J. J. Englander, D. B. Calhoun, and S. W. Englander, (1979) Analytical Biochemistry, 92, 517-524). Isotopic exchange data are presented as plots of X (the fraction of unexchanged peptide hydrogen atoms at time t) versus log(kot), where ko is the pH dependent rate constant for peptide groups exposed to the solvent. In the range of pD 2.25-6.9, at 20 degrees C, BTCI shows a continuous exchange curve which indicates that the exchange mechanism is of the EX2 type and no detectable conformational changes occur in the protein. Deviations from this exchange curve are found at pD 7.3 and 8.0. About 60% of the peptide hydrogens of BTCI are exchanged for delta Go less than or equal to 2 kcal/mol, and 90% for delta Go less than 6 kcal/mole. For reduced and carboxymethylated BTCI, exchange data suggest a much more open conformation in comparison with the unmodified protein. However, some residual structure appears to be maintained, after scission of the disulfide bonds. The exchange data indicate that, as a consequence of the formation of the beta-trypsin-BTCI complex, part of the peptide groups of the enzyme and/or inhibitor become less accessible to the isotopic exchange.
D003903:Deuterium / Q000378:metabolism*; D006859:Hydrogen / Q000378:metabolism*; D007474:Ion Exchange; D011487:Protein Conformation; D013056:Spectrophotometry, Ultraviolet; D014357:Trypsin / Q000378:metabolism; D014361:Trypsin Inhibitors / Q000378:metabolism*
D014361:Trypsin Inhibitors; D006859:Hydrogen; D003903:Deuterium; D014357:Trypsin