Shiro Takagi

Enzymic degradation of water-soluble dextran from Leuconostoc mesenteroides NRRL B-1299

Abstract Structural studies on the water-soluble dextran elaborated by Leuconostoc mesenteroides NRRL B-1299 gave several important results that not only supported previous results but afforded an insight into the association of average repeating-units in the whole molecule. Sequential degradation of soluble dextran from its nonreducing terminals was achieved with two different enzymes, namely, α- d -(1→2)-debranching enzyme and d -glucodextranase. The debranching enzyme removed four separate residues of α-(1→2)-branched d -glucose from the average repeating unit consisting of 15 d -glucosyl residues. d -Glucodextranase continuously produced d -glucose from the nonreducing terminals by an exo type of action, but internal branches greatly restricted its action. Extensive digestion of soluble dextran B-1299 with the two enzymes released 74.3% of d -glucose and gave a limit dextrin of high molecular weight containing 8.4% of 2,6-di- O -substituted d -glucosyl residues. 13 C-N.m.r. studies indicated a characteristic pattern of the α- d -(1→2)-branched structure, which was significantly changed on treatment with the debranching enzyme. Moreover, an ∼8-fold increase in the degree of linearity was observed after action of the debranching enzyme. The possible structure of water-soluble dextran B-1299 is discussed, based on a comparison of the limit dextrin with the native dextran, in regard to chemical structure, molecular-weight distribution, and degree of hydrolysis with two exo-enzymes. The native dextran might be constructed with at least ∼8,200 “twigs” of repeating unit, and there are 14 steps of connected twigs between the reducing and nonreducing terminals. Upon consecutive hydrolysis with the two exo-enzymes, most of the twigs located at the 14th step ( i.e., nonreducing terminals) were hydrolyzed to d -glucose.

An isomaltotriose-producing dextranase from Flavobacterium sp. M-73: Action pattern of the enzyme

Abstract An isomaltotriose-producing dextranase from Flavobacterium sp. M-73 (dextranase II) effectively hydrolyzed virtually linear dextrans, such as clinical dextran, dextrans T-10, T-110, or B-512F native dextran, to degrees ranging from 25 to 32%, expressed as apparent conversion into d -glucose. Highly branched dextrans, such as those from strais B-1298, B-1299, B-1307, or B-1416, were not good substrates for this enzyme. Studies on the action pattern of dextranase II on linear dextrans showed that this dextranase hydrolyzed high-molecular-weight substrates in an endo-type fashion. The major hydrolysis product was isomaltotriose. When isomalto-oligosaccharides were used as substrates, however, dextranase II hydrolyzed the low-molecular-weight substrates less effectively. Kinetic studies of the enzyme, using substrates of various chain lengths, showed that the affinity of the enzyme increased with increase of the molecular size of the substrate. Simultaneous use of the dextran α-(1→2)-debranching enzyme (dextranase I) with dextranase II greatly increased the extent of hydrolysis of branched dextrans from strains B-1298 and B-1299.