title: Catalytic properties of human Lys77-plasmin. A comparative steady-state and pre-steady-state study.
abstract: Values of kinetic parameters for the hydrolysis of esters and p-nitroanilides of L-lysine and L-arginine catalyzed by the Lys77 form of human plasmin (EC 3.4.21.7) have been determined between pH 5.5 and 8 (I = 0.1 M) at 21 +/- 0.5 degrees C. Over the whole pH range explored, Lys77-plasmin catalysis conforms to simple Michaelis-Menten kinetics, and steady-state and pre-steady-state data may be consistently fitted to the minimum three-step mechanism: E + S in equilibrium (k+1/k-1)E X S----(k+2)E X P + P1----(k+3)E + P2 In spite of the higher specificity of lysyl derivatives for Lys77-plasmin rather than the arginyl ones, kinetic parameters also depend on the nature of the N-alpha substituent and/or of the alcoholic or p-nitroanilidic moiety of the substrate. Among the esters and anilides considered, ZLysONp shows the most favourable kinetic parameters and may be the substrate of choice of Lys77-plasmin, in that it allows the determination of the enzyme concentration as low as 2 X 10(-9) M (about 1 X 10(-3) CU/ml), at the optimum pH value (approx. 8). Between pH 5.5 and 8, the pH profiles of kcat and kcat/Km for the Lys77-plasmin-catalyzed hydrolysis of ZLysONp and ZArgONp reflect the ionization of a single group (probably His-602 involved in the active site) with pKa values ranging between 6.4 and 6.6; at variance, values of Km are pH-independent.
authors.forename: P
authors.initials: P
authors.lastname: Ascenzi
authors.forename: A
authors.initials: A
authors.lastname: Torroni
authors.forename: E
authors.initials: E
authors.lastname: Menegatti
authors.forename: M
authors.initials: M
authors.lastname: Guarneri
authors.forename: G
authors.initials: G
authors.lastname: Amiconi
mesh_terms: D002384:Catalysis; D005341:Fibrinolysin / Q000494:pharmacology*; D006801:Humans; D006863:Hydrogen-Ion Concentration; D006868:Hydrolysis; D007700:Kinetics; D008239:Lysine; D010446:Peptide Fragments / Q000494:pharmacology*; D013379:Substrate Specificity
chemical_list: D010446:Peptide Fragments; C033809:lys-plasmin; D005341:Fibrinolysin; D008239:Lysine