Isolation of the membrane-bound 26 000-Mr penicillin-binding protein of Streptomyces strain K15 in the form of a penicillin-sensitive D-alanyl-D-alanine-cleaving transpeptidase

Nguyen-Distèche, Martine; Leyh-Bouille, Mélina; Ghuysen, Jean-Marie

Reference : Isolation of the membrane-bound 26 000-Mr penicillin-binding protein of Streptomyces ...


	Document type :	Scientific journals : Article
	Discipline(s) :	Life sciences : Biochemistry, biophysics & molecular biology
	To cite this reference:	http://hdl.handle.net/2268/83022

Title :	Isolation of the membrane-bound 26 000-Mr penicillin-binding protein of Streptomyces strain K15 in the form of a penicillin-sensitive D-alanyl-D-alanine-cleaving transpeptidase
Language :	English
Author, co-author :	Nguyen-Distèche, Martine [Université de Liège - ULiège > Faculté de Médecine, Institut de Chimie > Service de Microbiologie > >]
	Leyh-Bouille, Mélina [Université de Liège - ULiège > Faculté de Médecine, Institut de Chimie > Service de Microbiologie > > >]
	Ghuysen, Jean-Marie [Université de Liège - ULiège > Faculté de Médecine, Institut de Chimie > Service de Microbiologie > > >]
Publication date :	1-Oct-1982
Journal title :	Biochemical Journal
Publisher :	Portland Press
Volume :	207
Issue/season :	1
Pages :	109-115
Peer reviewed :	Yes (verified by ORBi)
Audience :	International
ISSN :	0264-6021
e-ISSN :	1470-8728
City :	London
Country :	United Kingdom
Keywords :	[en] bacterial proteins ; carboxypeptidases/isolation & purification ; carrier proteins/isolation & purification/metabolism ; cell membrane/enzymology ; cetrimonium compounds ; chromatography, affinity ; chromatography, gel ; detergents ; electrophoresis, polyacrylamide gel ; hexosyltransferases ; muramoylpentapeptide carboxypeptidase/antagonists & inhibitors/isolation ; purification/metabolism ; penicillin g/pharmacology ; penicillin-binding proteins ; peptidyl transferases ; streptomyces/*enzymology
Abstract :	[en] The membrane-bound, 26 000-Mr penicillin-binding protein of Streptomyces K15 has been isolated in the form of an effective, penicillin-sensitive D-alanyl-D-alanine-cleaving peptidase exhibiting high transpeptidase activity (greater than 95%) and very low carboxy-peptidase activity (less than 5%). The penicillin-binding protein/transpeptidase can be extracted directly from the mycelium with N-cetyl-NNN-trimethylammonium bromide (Cetavlon) and subsequently obtained at 90% purity and with an 8000-fold specific enrichment (when compared with the activity of the isolated membranes) by a two-step procedure involving Sephadex filtration and affinity chromatography on ampicillin-linked CH Sepharose 4B in the presence of detergent. At saturating concentrations of the co-substrates diacetyl-L-Lys-D-Ala-D-Ala and Gly-Gly, the catalytic-centre activity is about 0.3 s-1.
Funders :	Fonds de la Recherche Scientifique Médicale - FRSM ; National Institutes of Health - NIH
Target :	Researchers ; Professionals
Permalink :	http://hdl.handle.net/2268/83022

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