RESUMO
Chlorophenols comprise a major class of environmental contaminants. They are extensively used as insecticides, fungicides, mold inhibitors, antiseptics and disinfectants. We found some of these compounds to be strong inhibitors of zeta-crystallin. This oxidoreductase enzyme was isolated from camel lens and its enzymatic activity was inhibited by the chlorophenols tested in a time-independent but concentration-dependent manner. 2,4,5-Trichlorophenol was the most potent inhibitor (IC50 = 3 microM; Ki = 3.2 microM) whereas 4-chlorophenol was the least potent (IC50 = 4.1 mM). There appeared to be a relationship between the degree of chlorination of the phenols and inhibition of zeta-crystallin activity. The position of the chlorine substituent on the phenol may also influence the potency of these compounds.
Assuntos
Clorofenóis/farmacologia , Cristalinas/antagonistas & inibidores , Inibidores Enzimáticos/farmacologia , Cristalino/enzimologia , NAD(P)H Desidrogenase (Quinona)/antagonistas & inibidores , Animais , Camelus , Cristalinas/isolamento & purificação , Cristalinas/metabolismo , Cinética , Quinonas/metabolismo , Relação Estrutura-Atividade , zeta-CristalinasRESUMO
Camel lens zeta-crystallin/NADPH:quinone oxidoreductase activity was inhibited by chloranilic acid (2,5-dichloro-3,6-dihydroxy-1,4-benzoquinone) with NADPH as an electron donor and 9,10-phenanthrenequinone (PQ) as an electron acceptor in a time-independent but concentration dependent manner. The IC50 of chloranilic acid was 1 microM. The inhibition was noncompetitive with respect to both NADPH and PQ as deduced by Lineweaver-Burk plots. The estimated inhibition constant (Ki) was 0.8 microM for both NADPH and PQ. Examination of other benzoquinones suggested that the presence of -OH and -Cl on benzoquinone was essential for the inhibition.
Assuntos
Benzoquinonas/farmacologia , Cristalinas/antagonistas & inibidores , NAD(P)H Desidrogenase (Quinona)/antagonistas & inibidores , Animais , Camelus , Cloranila/farmacologia , CinéticaRESUMO
Kallikrein was isolated from bovine pancreatic tissue by a sequential procedure of anion-exchange, hydrophobic interaction and gel filtration chromatographies. A monospecific polyclonal antiserum to the pure enzyme was raised in rabbits and was used to set up a specific radioimmunoassay for bovine tissue kallikrein.
Assuntos
Calicreínas/isolamento & purificação , Pâncreas/enzimologia , Radioimunoensaio/métodos , Animais , Bovinos , Cromatografia em Agarose , Cromatografia em Gel , Cromatografia por Troca Iônica , Calicreínas/análise , Radioimunoensaio/normasRESUMO
Numerous biochemical properties (e.g. Mr, carbohydrate content, pI) were determined for kallikrein isolated from rat submandibular glands by a simple, rapid purification procedure. The kinetic behaviour of the enzyme towards various inhibitors and synthetic substrates was investigated. The effects of different salts and detergents on the esterolytic activity of the rat tissue kallikrein were recorded.
Assuntos
Calicreínas/isolamento & purificação , Glândula Submandibular/enzimologia , Animais , Arginina/análogos & derivados , Detergentes/farmacologia , Concentração de Íons de Hidrogênio , Técnicas In Vitro , Calicreínas/antagonistas & inibidores , Calicreínas/metabolismo , Cinética , Masculino , Oligopeptídeos , Ratos , Sais/farmacologia , Especificidade por Substrato , TemperaturaRESUMO
A prokallikrein was isolated from bovine pancreas by a multi-step procedure involving gel filtration, hydrophobic interaction and anion-exchange chromatographies. The purification was initially monitored by measurement of the kinin-releasing activity of the activated zymogen. Later, when the pure prokallikrein had been isolated, a specific radioimmunoassay for the zymogen was set up and that was employed to provide estimates of 323-fold and 28% for the overall degree of purification and percentage recovery of prokallikrein. The relative molecular weight of prokallikrein was found to be 26,900 by SDS gel electrophoresis and its isoelectric point was established as pH 4.55.
