Glycosidase induction in Pseudomonas stutzeri and properties of one of its amylases maltotetraohydrolase

Abstract

Pseudomonas stutzeri has been shown to produce extracellular amylases of different specificity, as well as a carbohydrase which hydrolyzed oligosaccharide but not starch. Pseudomonas stutzeri NCIB 11359 and NRRL B-3389 were adapted to, and then cultured in five different growth media. Both culture collections showed similar patterns in cell morphology and glucosidase secretion. Maltotetraohydrolase was produced in several of the media, a 'typical' alpha-amylase was exclusively produced in one of the defined media, and no glucanase in yet another medium. The endo-a-amylase which produced primarily maltose was present during early exponential cell growth and no longer detected by late exponential growth. When maltotetraohydrolase was produced, it was found throughout exponential growth, and in one of the media remained at maximum levels of activity into stationary phase. Maltotetraohydrolase hydrolyzed a suspension of commercial amylose in water producing 71-99% maltotetraose in the oligosaccharide fraction. Studies with maltotetraohydrolase demonstrated endo-amylase activity with the substrates blue amylose and with several 0-amylase limit dextrins. A limit dextrin was produced which could be further hydrolyzed by a-amylase. 1H-nmr studies showed that no a-1,6-glycosidic linkages were hydrolyzed and that the a-anomer was produced from the hydrolysis of a-1,4-linkages. Between 18-24% of the glycosidic linkages in amylopectin and 11-20% in B-limit dextrin were hydrolyzed. Maltotetraose was the primary product from hydrolysis of glucans but from the B-limit dextrins a high proportion of the smaller oligosaccharides was produced. Maltotetraohydrolase limit dextrin representing 38-54% of wheat amylopectin was purified and it had a molecular weight of 7000-15000; a degree of branching of 15%; ca. 13 branches per molecule; and an average chain length of 7. HPLC showed it to have a narrow weight distribution. Comparison of wheat and potato amylopectin indicated potato amylopectin to have more evenly distributed branching and probably less densely branched clusters.