Differential effect of halide anions on the hydrolysis of different dansyl substrates by thermolysin.
Biochemistry
; 33(21): 6516-23, 1994 May 31.
Article
en En
| MEDLINE
| ID: mdl-8204586
The effect of sodium halide salts on the hydrolysis of three of the dansyl (Dns) peptide substrates described in the previous paper (Yang & Van Wart, 1994) by thermolysin have been studied. Increasing concentrations of sodium chloride decrease the KM value for the hydrolysis of the tripeptides Dns-Gly-Phe-Ala and Dns-Ala-Phe-Ala but leave kcat unaltered. This kinetic behavior is described by a nonessential activation mechanism in which chloride binds preferentially to the enzyme-substrate complex. Similar trends are found for the sodium bromide and fluoride salts. In contrast, sodium chloride decreases both KM and kcat almost equally for the hydrolysis of Dns-Ala-Ala-Phe-Ala, leaving kcat/KM unchanged. Thus, chloride is an uncompetitive inhibitor of this substrate. Molecular modeling studies have been carried out in order to explain the effect of chloride on the binding of these dansyl peptides. The decrease in KM for the hydrolysis of all three substrates is attributed to an interaction of chloride with Arg-203 located in the active site to stabilize the enzyme-substrate complexes. The differential effect of chloride on the kcat values for the hydrolysis of the dansyl tripeptides vs dansyl tetrapeptide is related to differences in binding on the Pn side of the substrates. The tripeptides are predicted to bind to the active site of thermolysin in a single low-energy conformation. However, there are two populations of low-energy binding modes for the tetrapeptide, one of which is believed to be a more productive binding mode.(ABSTRACT TRUNCATED AT 250 WORDS)
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Colección:
01-internacional
Base de datos:
MEDLINE
Asunto principal:
Péptidos
/
Termolisina
/
Compuestos de Dansilo
/
Aniones
Tipo de estudio:
Prognostic_studies
Idioma:
En
Revista:
Biochemistry
Año:
1994
Tipo del documento:
Article
Pais de publicación:
Estados Unidos