Purification and characterization of a new kappa-carrageenase from a marine Cytophaga-like bacterium.
A bacterial strain able to degrade various sulfated galactans (carrageenans and agar) was isolated from the marine red alga Delesseria sanguinea. From the cell-free supernatant of cultures grown on crude lambda-carrageenan, a kappa-carrageenase was purified by ammonium sulfate fractionation, gel filtration on Sephacryl S 200 HR and ion-exchange chromatography on DEAE--Sepharose-CL6B. The purified kappa-carrageenase was detected as a single protein upon SDS/PAGE. Its molecular mass was estimated at 40 kDa. Activity was observed against kappa-carrageenan over the pH range 5.0-8.5 and was optimal at pH 7.2 in Tris buffer or 7.0 in Mops buffer. The enzyme activity remained stable at 30 degrees C, but only for up to 1 h at 40 degrees C. Analysis of the degradation products of the kappa-carrageenase by gel filtration and 13C-NMR spectroscopy indicated that the enzyme degrades kappa-carrageenan down to the level of the kappa-neocarratetraose sulfate. The properties of this new enzyme are compared with those of previously characterized carrageenases.
Potin
P
P
Station Biologique, CNRS UPR 4601, Roscoff, France.
Sanseau
A
A
Le Gall
Y
Y
Rochas
C
C
Kloareg
B
B
D001426:Bacterial Proteins; D002351:Carrageenan / Q000378:metabolism; D002845:Chromatography; D006026:Glycoside Hydrolases / Q000302:isolation & purification* / Q000378:metabolism; D006868:Hydrolysis; D007700:Kinetics; D009682:Magnetic Resonance Spectroscopy; D008970:Molecular Weight; D000461:Rhodophyta / Q000201:enzymology*; D013379:Substrate Specificity
D001426:Bacterial Proteins; D002351:Carrageenan; D006026:Glycoside Hydrolases; C019846:kappa-carrageenase protein, Alteromonadaceae