A Novel Bacillus Pullulanase-Its Properties and Application in the Glucose Syrups Industry

抄録

The majority of starches used in the manu-facture of glucose syrups contain 75-85% amylopectin.1) Amylopectin is a highly branched polysaccharide consisting of linear chains of 1, 4-α-linked D-glucose residues, joined together by 1, 6-α-glucosidic linkages. The branch points occur on average every 20-25 D-glucose units, so that amylopectin contains 4-5% of 1, 6-α-glucosidic linkages.^2-4) The 1, 6-α-glucosidic linkages act as a kind of barrier to the action of exo-acting, saccharif ying amylases such as glucoamylases or maltogenic /3-amylases. Endo-acting α-amylases are able to by-pass the branch points, ⁵⁾ but in general are not capable cf hydrolyzing the 1, 6-α-glucosidic linkage. Recent work by Kobayashi and co-workers has shown that at least one α-amylase (from Thermoactinomyces vulgaris) can hydro-lyze 1, 6-α-glucosidic linkages, in addition to 1, 4-α-glucosidic linkages.⁶⁾ Glucoamylases can slowly hydrolyze 1, 6-α-glucosidic linkages in amylopectin and partially hydrolyzed amylopectin, ⁷⁾ but the action of maltogenic exo-amylases ceases as a branch point is approached.⁸⁾ It is therefore obvious that the efficiency of the saccharification reaction could be improved by incorporating a specific amylopectin debranchina enzyme in the system. Debranching enzyme such as isoamylase [EC 3.2.1.68, glycogen 6-glucanohydrolase] and pullulanase [EC 3.2.1.41, pullulan 6-glucanohy-drolase] have been known for many years, ⁹⁾ but their use in the glucose syrups industry is far from widespread. Pullulanases from Klebsiella pneumoniae, ^{10, 11)} Streptomyces sp., ¹²⁾ and Bacillus cereus var, mycoides¹³⁾ and isoamylases from Pseudomonas amyloderamosa^{14, 15)} and Cy-tophaga sp.¹⁶⁾ are not sufficiently thermostable to be used at 60°C. Moreover the Pseudomonas isoamylase was the only debranching enzyme sufficiently acidophilic to be used at a pH of around 4. 5.¹⁷, ¹⁸⁾ After an extensive screening programme, our research laboratories succeeded in isolating a species of Bacillus which produced a thermostable, acidophilic pullulanase which was free from glucoamylase, β-amylase and a-amylase side activities. Some of the properties of this enzyme will now be described.