title: Kinetic peculiarities of human tissue kallikrein: 1--substrate activation in the catalyzed hydrolysis of H-D-valyl-L-leucyl-L-arginine 4-nitroanilide and H-D-valyl-L-leucyl-L-lysine 4-nitroanilide; 2--substrate inhibition in the catalyzed hydrolysis of N alpha-p-tosyl-L-arginine methyl ester.
abstract: Hydrolysis of D-valyl-L-leucyl-L-lysine 4-nitroanilide (1), D-valyl-L-leucyl-L-arginine 4-nitroanilide (2), and N alpha-p-tosyl-L-arginine methyl ester (3) by human tissue kallikrein was studied throughout a wide range of substrate concentrations. At low substrate concentrations, the hydrolysis followed Michaelis-Menten kinetics but, at higher substrate concentrations, a deviation from Michaelis-Menten behavior was observed. With the nitroanilides, a significant increase in hydrolysis rates was observed, while with the ester, a significant decrease in hydrolysis rates was observed. The results for substrates (1) and (3) can be accounted for by a model based on the hypothesis that a second substrate molecule binds to the ES complex to produce a more active or an inactive SES complex. The deviation observed for substrate (2) can be explained as a bimolecular reaction between the enzyme-substrate complex and a free substrate molecule.
authors.affiliation: Departamento de Análises Clínicas e Toxicológicas, Faculdade de Farmácia, Universidade Federal de Minas Gerais, 30123-970 Belo Horizonte, MG, Brazil.
authors.forename: Marinez O
authors.initials: MO
authors.lastname: Sousa
authors.forename: Tânia L S
authors.initials: TL
authors.lastname: Miranda
authors.forename: Caroline N
authors.initials: CN
authors.lastname: Maia
authors.forename: Eustáquio R
authors.initials: ER
authors.lastname: Bittar
authors.forename: Marcelo M
authors.initials: MM
authors.lastname: Santoro
authors.forename: Amintas F S
authors.initials: AF
authors.lastname: Figueiredo
mesh_terms: D002384:Catalysis; D002863:Chromogenic Compounds / Q000494:pharmacology; D006801:Humans; D006868:Hydrolysis; D007610:Kallikreins / Q000737:chemistry* / Q000378:metabolism*; D007700:Kinetics; D008956:Models, Chemical; D009842:Oligopeptides / Q000494:pharmacology; D011485:Protein Binding; D013379:Substrate Specificity; D014106:Tosylarginine Methyl Ester / Q000737:chemistry
chemical_list: D002863:Chromogenic Compounds; D009842:Oligopeptides; C018788:valyl-leucyl-lysine 4-nitroanilide; C031006:Val-Leu-Arg-p-nitroanilide; D014106:Tosylarginine Methyl Ester; D007610:Kallikreins