ORBi: Chavagnat Frederic - Cloning, sequencing and overexpression in Escherichia coli of the alginatelyase-encoding aly gene of Pseudomonas alginovora: identification of three classes of alginate lyases

Reference : Cloning, sequencing and overexpression in Escherichia coli of the alginatelyase-encoding...


	Document type :	Scientific journals : Article
	Discipline(s) :	Life sciences : Biochemistry, biophysics & molecular biology
	To cite this reference:	http://hdl.handle.net/2268/90900

Title :	Cloning, sequencing and overexpression in Escherichia coli of the alginatelyase-encoding aly gene of Pseudomonas alginovora: identification of three classes of alginate lyases
Language :	English
Author, co-author :	Chavagnat, Frederic [Université Claude Bernard - Lyon 1 - UCLB > > Laboratoire de Biochimie Analytique et Synthèse Bioorganique > >]
	Duez, Colette [Université de Liège - ULg > > Centre d'Ingénierie des Protéines > > >]
	Guinand, Micheline [Université Claude Bernard - Lyon 1 - UCLB > > Laboratoire de Biochimie Analytique et Synthèse Bioorganique > > >]
	Potin, Philippe [Station Biologique, Roscoﬀ (France) > > Centre d’Etudes d’Océanographie et de Biologie Marine > >]
	Barbeyron, Tristan [Station Biologique, Roscoﬀ (France) > > Centre d’Etudes d’Océanographie et de Biologie Marine > >]
	Henrissat, Bernard [Centre de Recherches sur les Macromolécules Végétales (CNRS) > > > >]
	Wallach, Jean [Université Claude Bernard - Lyon 1 - UCLB > > Laboratoire de Biochimie Analytique et Synthèse Bioorganique > >]
	Ghuysen, Jean-Marie [Université de Liège - ULg > > Centre d'Ingénierie des Protéines > >]
Publication date :	1996
Journal title :	Biochemical Journal
Publisher :	Portland Press
Volume :	319
Issue/season :	Pt 2
Pages :	575-583
Audience :	International
ISSN :	0264-6021
e-ISSN :	1470-8728
City :	London
Country :	United Kingdom
Keywords :	[en] amino acid sequence ; base sequence ; cloning, molecular ; escherichia coli/genetics ; gene transfer techniques ; molecular sequence data ; polysaccharide-lyases/genetics/metabolism ; pseudomonas/enzymology/genetics ; sequence alignment ; sequence analysis
Abstract :	[en] A gene of Pseudomonas alginovora, called aly, has been cloned in Escherichia coli using a battery of PCR techniques and sequenced. It encodes a 210-amino-acid alginate lyase (EC 4.2.2.3), Aly, in the form of a 233-amino-acid precursor. P. alginovora Aly has been overproduced in E. coli with a His-tag sequence fused at the C-terminal end under conditions in which the formation of inclusion bodies is avoided. His-tagged P. alginovora Aly has the same enzymic properties as the wild-type enzyme and has the specificity of a mannuronate lyase. It can be purified in a one-step procedure by affinity chromatography on Ni(2+)-nitriloacetate resin. The yield is of 5 mg of enzyme per litre of culture. The amplification factor is 12.5 compared with the level of production by wild-type P. alginovora. The six alginate lyases of known primary structure fall into three distinct classes, one of which comprises the pair P. alginovora Aly and Klebsiella pneumoniae Aly.
Research units :	Centre d'Ingénierie des Protéines - CIP
Funders :	Fonds de la Recherche Scientifique (Communauté française de Belgique) - F.R.S.-FNRS ; Fonds de la Recherche Scientifique Médicale - FRSM
Target :	Researchers ; Professionals
Permalink :	http://hdl.handle.net/2268/90900

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