Purification and properties of alternanase, a novel endo-alpha-1,3-alpha-1,6-D-glucanase.
A newly isolated soil bacterium strain NRRL B-21195, tentatively identified as a Bacillus species, was found to be a constitutive producer of a novel type of glycanase that hydrolyses in an endo-fashion the polysaccharide alternan, an alpha-1,3-alpha-1,6-D-glucan, referred to in the literature as B-1355 dextran (fraction S), synthesized from sucrose by alternansucrase of Leuconostoc mesenteroides. The glycanase, named alternanase, has been purified to homogeneity from a cell-free culture fluid of the bacillus grown in a liquid medium containing D-glucose, and has been characterized. The enzyme has a molecular mass of 110000 Da (SDS/PAGE) and an isoelectric point of approximately 4.0. Optimum activity occurs at pH 7 and at a temperature of 40 degrees C. The enzyme is stable up to 50 degrees C but loses activity rapidly at 60 degrees C. Its action is inhibited by EDTA and stimulated by Ca2+. The enzyme requires, for its action, D-glucan chains in which alpha-1,3-linkages alternate with alpha-1,6-linkages; i.e., it is specific for alternan. Monitoring of alternan hydrolysis by determination of liberated reducing sugars pointed to an unusually low extent of hydrolysis and a low specific activity of the enzyme. As shown in the accompanying paper [Côté, G. L. & Biely, P. (1994) Eur. J. Biochem. 226, 641-648] the reason for this finding is that the main hydrolytic products are non-reducing, novel types of cyclic oligosaccharides.
Biely
P
P
Institute of Chemistry, Slovak Academy of Sciences, Bratislava.
Côté
G L
GL
Burgess-Cassler
A
A
D001407:Bacillus / Q000201:enzymology*; D002118:Calcium / Q000494:pharmacology; D002236:Carbohydrate Conformation; D002412:Cations; D004492:Edetic Acid / Q000494:pharmacology; D004795:Enzyme Stability; D005936:Glucans / Q000737:chemistry / Q000378:metabolism; D006026:Glycoside Hydrolases / Q000737:chemistry / Q000302:isolation & purification* / Q000378:metabolism; D006863:Hydrogen-Ion Concentration; D007526:Isoelectric Point; D007700:Kinetics; D008970:Molecular Weight; D013379:Substrate Specificity; D013696:Temperature
D002412:Cations; D005936:Glucans; D004492:Edetic Acid; D006026:Glycoside Hydrolases; C091143:alternanase; D002118:Calcium