Microenvironmental effects on enzyme catalysis. A kinetic study of hydrophobic derivatives of chymotrypsin.
The aim of this work was to study the role of hydrophobic interactions in the enzymic activity of chymotrypsin. The amino groups of chymotrypsin were chemically modified by aliphatic aldehydes of various chain lengths - acetaldehyde, butyraldehyde, hexanal - and with two aldehydes of different steric hindrance - benzaldehyde and trimethyl acetaldehyde. After a rapid study of the derivated enzymes, the hexylchymotrypsin has been chosen for its new catalytic properties: the Michaelis constant is not modified and the maximal velocity with N-glutaryl-L-phenylalanine-4-nitroaniline is increased to 164%. The increase is due to the increase of the acylation constant, k2, by 230%. The value of k3 is not modified or less modified. In the modified enzyme, 85% of free amino acids are still able to react with trinitrobenzenesulphonic acid. The optimum pH is shifted by one pH unit towards the alkaline pH. The thermodynamic study shows that the catalytic process itself is not modified. The increase in Vm could be a simple increase of k2 linked to a modification of the site or of the protein. The phenomenon described is very specific and obtained only with one modification, hexanal, and with one enzyme, alpha-chymotrypsin.
D000818:Animals; D001665:Binding Sites; D002417:Cattle; D002918:Chymotrypsin / Q000378:metabolism*; D006358:Hot Temperature; D006863:Hydrogen-Ion Concentration; D066298:In Vitro Techniques; D007700:Kinetics; D011489:Protein Denaturation; D012995:Solubility; D013329:Structure-Activity Relationship; D013816:Thermodynamics
D002918:Chymotrypsin