title: Degradation of lambda-carrageenan by Pseudoalteromonas carrageenovora lambda-carrageenase: a new family of glycoside hydrolases unrelated to kappa- and iota-carrageenases.
abstract: Carrageenans are sulfated galactans found in the cell walls of red seaweeds. They are classified according to the number and the position of sulfate ester groups. lambda-Carrageenan is the most sulfated carrageenan and carries at least three sulfates per disaccharide unit. The sole known depolymerizing enzyme of lambda-carrageenan, the lambda-carrageenase from Pseudoalteromonas carrageenovora, has been purified, cloned and sequenced. Sequence analyses have revealed that the lambda-carrageenase, referred to as CglA, is the first member of a new family of GHs (glycoside hydrolases), which is unrelated to families GH16, that contains kappa-carrageenases, and GH82, that contains iota-carrageenases. This large enzyme (105 kDa) features a low-complexity region, suggesting the presence of a linker connecting at least two independent modules. The N-terminal region is predicted to fold as a beta-propeller. The main degradation products have been purified and characterized as neo-lambda-carratetraose [DP (degree of polymerization) 4] and neo-lambda-carrahexaose (DP6), indicating that CglA hydrolyses the beta-(1-->4) linkage of lambda-carrageenan. LC-MALLS (liquid chromatography-multi-angle laser light scattering) and (1)H-NMR monitoring of the enzymatic degradation of lambda-carrageenan indicate that CglA proceeds according to an endolytic mode of action and a mechanism of inversion of the anomeric configuration. Using 2-aminoacridone-labelled neo-lambda-carrabiose oligosaccharides, in the present study we demonstrate that the active site of CglA comprises at least 8 subsites (-4 to +4) and that a DP6 oligosaccharide binds in the subsites -4 to +2 and can be hydrolysed into DP4 and DP2.
authors.affiliation: Centre National de la Recherche Scientifique, Université Pierre et Marie Curie-Paris6, Unité Mixte de Recherche 7139 Marine Plants and Biomolecules, Station Biologique, F-29682 Roscoff Cedex, Bretagne, France.
authors.forename: Marion
authors.initials: M
authors.lastname: Guibet
authors.forename: Sébastien
authors.initials: S
authors.lastname: Colin
authors.forename: Tristan
authors.initials: T
authors.lastname: Barbeyron
authors.forename: Sabine
authors.initials: S
authors.lastname: Genicot
authors.forename: Bernard
authors.initials: B
authors.lastname: Kloareg
authors.forename: Gurvan
authors.initials: G
authors.lastname: Michel
authors.forename: William
authors.initials: W
authors.lastname: Helbert
mesh_terms: D001426:Bacterial Proteins / Q000378:metabolism*; D002236:Carbohydrate Conformation; D002351:Carrageenan / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism*; D002850:Chromatography, Gel; D006026:Glycoside Hydrolases / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism*; D007700:Kinetics; D009682:Magnetic Resonance Spectroscopy; D008969:Molecular Sequence Data; D009844:Oligosaccharides / Q000737:chemistry / Q000378:metabolism; D016366:Open Reading Frames; D031161:Pseudoalteromonas / Q000201:enzymology*; D011994:Recombinant Proteins / Q000737:chemistry / Q000378:metabolism; D013379:Substrate Specificity
chemical_list: D001426:Bacterial Proteins; D009844:Oligosaccharides; D011994:Recombinant Proteins; D002351:Carrageenan; D006026:Glycoside Hydrolases; C516835:lambda-carrageenase, Pseudoalteromonas