Catalogo Articoli (Spogli Riviste)

OPAC HELP

Titolo:
Slow-binding inhibition of gamma-glutamyl transpeptidase by gamma-boroGlu
Autore:
Stein, RL; DeCicco, C; Nelson, D; Thomas, B;
Indirizzi:
DuPont Pharmaceut Co, Dept Chem Enzymol, Expt Stn, Wilmington, DE 19880 USA DuPont Pharmaceut Co Wilmington DE USA 19880 tn, Wilmington, DE 19880 USA DuPont Pharmaceut Co, Dept Med Chem, Wilmington, DE 19880 USA DuPont Pharmaceut Co Wilmington DE USA 19880 em, Wilmington, DE 19880 USA
Titolo Testata:
BIOCHEMISTRY
fascicolo: 19, volume: 40, anno: 2001,
pagine: 5804 - 5811
SICI:
0006-2960(20010515)40:19<5804:SIOGTB>2.0.ZU;2-Q
Fonte:
ISI
Lingua:
ENG
Soggetto:
HUMAN-LEUKOCYTE ELASTASE; AMINO-ACIDS; BORONIC ACIDS; CHYMOTRYPSIN; DERIVATIVES; GLUTATHIONE; DIPEPTIDES; CATALYSIS;
Tipo documento:
Article
Natura:
Periodico
Settore Disciplinare:
Life Sciences
Citazioni:
21
Recensione:
Indirizzi per estratti:
Indirizzo: Stein, RL DuPont Pharmaceut Co, Dept Chem Enzymol, Expt Stn, E400-4462,Route 141 & Henry Clay Rd, Wilmington, DE 19880 USA DuPont Pharmaceut Co E400-4462,Route 141 & Henry Clay Rd Wilmington DE USA 19880
Citazione:
R.L. Stein et al., "Slow-binding inhibition of gamma-glutamyl transpeptidase by gamma-boroGlu", BIOCHEM, 40(19), 2001, pp. 5804-5811

Abstract

gamma -Glutamyl transpeptidase (gamma GTase) catalyzes the transfer of thegamma -glutamyl moiety of gamma -glutamyl-derived peptides, such as glutathione (gamma Glu-Cys-Gly), and anilides, such as gamma -glutamyl-7-amido-4-methylcoumarin (gamma Glu-AMC), to acceptor molecules, including water and various dipeptides. These acyl-transfer reactions all occur through a common acyl-enzyme intermediate formed from attack of an active site hydroxyl onthe gamma -carbonyl carbon of gamma Glu-X with displacement of X. In this paper, we report that gamma GTase is potently inhibited by the gamma -boronic acid analogue of L-glutamic acid, 3-amino-3-carboxypropaneboronic acid (gamma -boroGlu). We propose that gamma -boroGlu adds to the active site hydroxyl of gamma GTase to form a covalent, tetrahedral adduct that resembles tetrahedral transition states and intermediates that occur along the reaction pathway for gamma GTase-catalyzed reactions. Our studies demonstrate that gamma -boroGlu is a competitive inhibitor of the gamma GTase-catalyzed hydrolysis of gamma Glu-AMC with a K-i value of 35 nM. Kinetics of inhibitionstudies allow us to estimate the following values: k(on) = 400 mM(-1) s(-1) and k(off) = 0.02 s(-1). We also found that gamma -boroGlu is an uncompetitive inhibitor of Gly-Gly-promoted transamidation of gamma Glu-AMC. This observation is consistent with the kinetic mechanism we determined for gammaGTase-catalyzed transamidation of gamma Glu-AMC by Gly-Gly to form gamma Glu-Gly-Gly. To probe rate-limiting transition states for gamma GTase catalysis and inhibition, we determined solvent deuterium isotope effects. Solvent isotope effects on k(c)/K-m for hydrolysis of gamma Glu-AMC and k(on) forinhibition by gamma -boroGlu are identical and equal unity, suggesting that the processes governed by these rate constants are both rate-limited by astep that is insensitive to solvent deuterium such as a conformational fluctuation of the initially formed E-S or E-I complex. In contrast, the solvent isotope effect on k(c) is 2.4. k(c) is rate-limited by hydrolysis of theacyl-enzyme intermediate that is formed during reaction of gamma GTase with gamma Glu-AMC. Thus, the magnitude of this isotope effect suggests the formation of a catalytically important protonic bridge in the rate-limiting transition state for deacylation.

ASDD Area Sistemi Dipartimentali e Documentali, Università di Bologna, Catalogo delle riviste ed altri periodici
Documento generato il 27/10/20 alle ore 11:13:31