1887

Abstract

Extracellular endo-(1→3)-1-glucanase (EC 3.2.1.59) was optimally induced in WL-12 when grown in a mineral medium containing intact cells of or extensively purified (1→3)-α-glucan as the carbon source. The enzyme activity was separated from most other proteins, including the -glucanases, by affinity adsorption of the enzyme on water-insoluble colloidal (1→3)-α-glucan. The enzyme was released by allowing it to hydrolyse this substrate and purified further by DEAE-agarose chromatography and polyacrylamide P-150 gel filtration. Passage over DEAE-agarose apparently also separated a minor (1→3)-α-glucanase component. The principal (1→3)-α-glucanase was specific for the (1→3)-α-glucosidic bond and hydrolysed (1→3)-α-glucan endolytically to a mixture of nigerose and glucose with a transient accumulation of nigerotetraose. The reaction proceeded at a constantly declining rate with either colloidal (1→3)-α-glucan or with the soluble carboxymethyl-(1→3)-α-glucan as substrate. Nigeran, containing alternating (1→3)-α- and (1→4)-α-linkages, was not hydrolysed. Substrate dependence showed Michaelis-Menten kinetics. There was a pH optimum of 7·5 to 8·5 with a pronounced shoulder at pH 5 to 7. The molecular weight was estimated by slab gel electrophoresis in sodium dodecyl sulphate to be 134000. The enzyme did not appear to require divalent metal ions because its activity was stimulated by EDTA. There was evidence for essential thiol groups. Carbohydrate was not detected in the enzyme.

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1980-05-01
2021-05-16
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