Selective oxidation of Met-192 in bovine alpha-chymotrypsin. Effect on catalytic and inhibitor binding properties.
Catalytic and inhibitor binding properties of bovine alpha-chymotrypsin, in which the Met-192 residue has been converted by treatment with chloramine T to the sulfoxide derivative (Met(O)192 alpha-chymotrypsin), have been examined relative to the native enzyme (alpha-chymotrypsin), between pH 4.5 and 8.0 (mu = 0.1), and/or 5.0 degrees C and 40.0 degrees C. Values of kcat, k+2 and/or k+3 for the hydrolysis of all the substrates examined (i.e., tMetAcONp, ZAlaONp, ZLeuONp, ZLysONp and ZTyrONp) catalyzed by native and Met(O)192 alpha-chymotrypsin are similar, as well as values of Km for the hydrolysis of ZLeuONp, ZLysONp and ZTyrONp. On the other hand, Ks and Km values for the hydrolysis of ZAlaONp and tMetAcONp are decreased by about 5-fold. Met-192 oxidation does not affect the kinetic and thermodynamic parameters for the (de)stabilization of the complex formed between the proteinase and the bovine basic pancreatic trypsin inhibitor. On the other hand, the recognition process between between alpha-chymotrypsin and the recombinant proteinase inhibitor eglin c from the leech Hirudo medicinalis is influenced by the oxidation event. Considering known molecular models, the observed catalytic and inhibitor binding properties of native and Met(O)192 alpha-chymotrypsin were related to the inferred stereochemistry of the proteinase-substrate and proteinase-inhibitor contact region(s).
Cutruzzolá
F
F
C.N.R., Department of Biochemical Sciences Alessandro Rossi Fanelli, University of Rome La Sapienza, Italy.
Ascenzi
P
P
Barra
D
D
Bolognesi
M
M
Menegatti
E
E
Sarti
P
P
Schnebli
H P
HP
Tomova
S
S
Amiconi
G
G
D000818:Animals; D002384:Catalysis; D002417:Cattle; D002918:Chymotrypsin / Q000037:antagonists & inhibitors / Q000378:metabolism*; D006863:Hydrogen-Ion Concentration; D008715:Methionine / Q000378:metabolism*; D010084:Oxidation-Reduction; D013057:Spectrum Analysis; D013816:Thermodynamics
D008715:Methionine; D002918:Chymotrypsin