title: Catalytic properties and specificity of a recombinant, overexpressed D-mannuronate lyase.
abstract: Lysis of alginates and of their saturated and unsaturated fragments was monitored by 1H NMR spectroscopy. AlxM(B) alginate lyase performs beta-elimination on the mannuronic acid (M) residues. It does not cleave the guluronic acid (G) sequences, nor the M-G or the G-M diads. In consequence, it is a true mannuronate lyase. The end product of the reaction is O-(4-deoxy-alpha-L-ery-thro-hex-4-enopyranosyl-uronic acid)-(1->(4)-O-(beta-D-mannopyranosyluronic acid)-(1->4)-O-beta-D-mannpyranuronic acid. Viscosity measurements made during degradation of a polymannuronate alginate showed that AlxM(B) behaves as an endo-enzyme. HPLC analysis of the degradation products of oligomannuronates and oligoalginates suggested that the beta-elimination requires the interaction of the enzyme with at least three sequential mannuronic acid residues. The catalytic site may possess 5 sub-sites and accommodate pentamers with different M/G ratio. Kinetic measurements showed that the specificity constant Vm/Km increased with the number of mannuronic acid residues. AlxM(B) may be reversibly inhibited by heteropolymeric blocks in a competitive manner.
authors.affiliation: Laboratoire de Biochimie Analytique et Synthèse Bioorganique, Université Claude Bernard, Villeurbanne, France.
authors.forename: F
authors.initials: F
authors.lastname: Chavagnat
authors.forename: A
authors.initials: A
authors.lastname: Heyraud
authors.forename: P
authors.initials: P
authors.lastname: Colin-Morel
authors.forename: M
authors.initials: M
authors.lastname: Guinand
authors.forename: J
authors.initials: J
authors.lastname: Wallach
mesh_terms: D000464:Alginates / Q000378:metabolism; D007700:Kinetics; D019906:Nuclear Magnetic Resonance, Biomolecular; D011133:Polysaccharide-Lyases / Q000378:metabolism*; D011994:Recombinant Proteins / Q000378:metabolism; D013379:Substrate Specificity
chemical_list: D000464:Alginates; D011994:Recombinant Proteins; D011133:Polysaccharide-Lyases; C016158:poly(beta-D-mannuronate) lyase