DESCRIBE <http://purl.uniprot.org/citations/16108757>

Subject	Predicate	Object
http://purl.uniprot.org/citations/16108757	http://www.w3.org/1999/02/22-rdf-syntax-ns#type	http://purl.uniprot.org/core/Journal_Citation
http://purl.uniprot.org/citations/16108757	http://www.w3.org/1999/02/22-rdf-syntax-ns#type	http://purl.uniprot.org/core/Journal_Citation
http://purl.uniprot.org/citations/16108757	http://www.w3.org/2000/01/rdf-schema#comment	"cABC I (chondroitinase ABC I) from Proteus vulgaris is a GalAG (galactosaminoglycan) depolymerizing lyase that cleaves its substrates at the glycosidic bond via beta-elimination. cABC I cleaves a particularly broad range of GalAG substrates, including CS (chondroitin sulphate), DS (dermatan sulphate) and hyaluronic acid. We recently cloned and recombinantly expressed cABC I in Escherichia coli, and completed a preliminary biochemical characterization of the enzyme. In the present study, we have coupled site-directed mutagenesis of the recombinant cABC I with a structural model of the enzyme-substrate complex in order to investigate in detail the roles of active site amino acids in the catalytic action of the enzyme. The putative catalytic residues His-501, Tyr-508, Arg-560 and Glu-653 were probed systematically via mutagenesis. Assessment of these mutants in kinetic and end-point assays provided direct evidence on the catalytic roles of these active-site residues. The crystal structure of the native enzyme provided a framework for molecular docking of representative CS and DS substrates. This enabled us to construct recombinant enzyme-substrate structural complexes. These studies together provided structural insights into the effects of the mutations on the catalytic mechanism of cABC I and the differences in its processing of CS and DS substrates. All His-501 mutants were essentially inactive and thereby implicating this amino acid to play the critical role of proton abstraction during catalysis. The kinetic data for Glu-653 mutants indicated that it is involved in a hydrogen bonding network in the active site. The proximity of Tyr-508 to the glycosidic oxygen of the substrate at the site of cleavage suggested its potential role in protonating the leaving group. Arg-560 was proximal to the uronic acid C-5 proton, suggesting its possible role in the stabilization of the carbanion intermediate formed during catalysis."xsd:string
http://purl.uniprot.org/citations/16108757	http://purl.org/dc/terms/identifier	"doi:10.1042/bj20050532"xsd:string
http://purl.uniprot.org/citations/16108757	http://purl.org/dc/terms/identifier	"doi:10.1042/bj20050532"xsd:string
http://purl.uniprot.org/citations/16108757	http://purl.uniprot.org/core/author	"Pojasek K."xsd:string
http://purl.uniprot.org/citations/16108757	http://purl.uniprot.org/core/author	"Pojasek K."xsd:string
http://purl.uniprot.org/citations/16108757	http://purl.uniprot.org/core/author	"Raman R."xsd:string
http://purl.uniprot.org/citations/16108757	http://purl.uniprot.org/core/author	"Raman R."xsd:string
http://purl.uniprot.org/citations/16108757	http://purl.uniprot.org/core/author	"Sasisekharan R."xsd:string
http://purl.uniprot.org/citations/16108757	http://purl.uniprot.org/core/author	"Sasisekharan R."xsd:string
http://purl.uniprot.org/citations/16108757	http://purl.uniprot.org/core/author	"Bosques C.J."xsd:string
http://purl.uniprot.org/citations/16108757	http://purl.uniprot.org/core/author	"Bosques C.J."xsd:string
http://purl.uniprot.org/citations/16108757	http://purl.uniprot.org/core/author	"Capila I."xsd:string
http://purl.uniprot.org/citations/16108757	http://purl.uniprot.org/core/author	"Capila I."xsd:string
http://purl.uniprot.org/citations/16108757	http://purl.uniprot.org/core/author	"Prabhakar V."xsd:string
http://purl.uniprot.org/citations/16108757	http://purl.uniprot.org/core/author	"Prabhakar V."xsd:string
http://purl.uniprot.org/citations/16108757	http://purl.uniprot.org/core/date	"2005"xsd:gYear
http://purl.uniprot.org/citations/16108757	http://purl.uniprot.org/core/date	"2005"xsd:gYear
http://purl.uniprot.org/citations/16108757	http://purl.uniprot.org/core/name	"Biochem. J."xsd:string
http://purl.uniprot.org/citations/16108757	http://purl.uniprot.org/core/name	"Biochem. J."xsd:string
http://purl.uniprot.org/citations/16108757	http://purl.uniprot.org/core/pages	"395-405"xsd:string
http://purl.uniprot.org/citations/16108757	http://purl.uniprot.org/core/pages	"395-405"xsd:string
http://purl.uniprot.org/citations/16108757	http://purl.uniprot.org/core/title	"Biochemical characterization of the chondroitinase ABC I active site."xsd:string
http://purl.uniprot.org/citations/16108757	http://purl.uniprot.org/core/title	"Biochemical characterization of the chondroitinase ABC I active site."xsd:string