High 1,3-beta-glucanase activity was detected during autolysis in a culture medium containing Penicillium oxalicum. It was due to the combined action of four enzymes. The purification process for the major enzyme produced a homogeneous band in the SDS polyacrylamide gel that corresponded to a molecular weight of 79,400 daltons. The enzyme pI was 6.3 and it was only active against 1,3-beta-glucans, with a S0.5 of 0.23 mg ml-1 against laminarin. The enzymatic optima were found at pH 4 and 55 degrees C, and instability was evident when pH and temperature were altered. The enzyme was not active against oxidated laminarin and was barely inhibited by glucono-D-lactone. Hg2+, Ag+ and Fe2+ were effective inhibitors. The enzyme was adsorbed by concanavalin-A-sepharose.
A systematic study on autolysis of the cell walls of fungi has been made on Neurospora crassa, Botrytis cinerea, Polystictus versicolor, Aspergillus nidulans, Schizophyllum commune, Aspergillus niger, and Mucor mucedo. During autolysis each fungus produces the necessary lytic enzymes for its autodegradation. From autolyzed cultures of each fungus enzymatic precipitates were obtained. The degree of lysis of the cell walls, obtained from non-autolyzed mycelia, was studied by incubating these cell walls with and without a supply of their own lytic enzymes. The degree of lysis increased with the incubation time and generally was higher with a supply of lytic enzymes. Cell walls from mycelia of different ages were obtained. A higher degree of lysis was always found, in young cell walls than in older cell walls, when exogenous lytic enzymes were present. In all the fungi studied, there is lysis of the cell walls during autolysis. This is confirmed by the change of the cell wall structure as well as by the degree of lysis reached by the cell wall and the release of substances, principally glucose and N-acetylglucosamine in the medium.
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The biological upgrading of wheat straw with Streptomyces cyaneus was examined through the analysis of chemical and structural changes of the transformed substrate during solid-state fermentation. Analysis of enzymes produced during the growth of S. cyaneus showed that phenol oxidase was the predominant enzyme. The reduction in Klason lignin content (16.4%) in the transformed substrate indicated the ability of this strain to delignify lignocellulose residues and suggests a role for phenol oxidase in the bacterial delignification process. Microscopic examination of the transformed substrate showed that the initial attack occurred at the less lignified cell walls (phloem and parenchyma), while xylem and sclerenchyma were slowly degraded. The pattern of degradation of sclerenchymatic tissues by S. cyaneus showed delamination between primary and secondary walls and between S1 and S2 due to partial removal of lignin. In the later stages of the decay a disorganization of the secondary walls was detected on account of fibrillation of this layer. A comparison of the properties of the pulp from wheat straw transformed by S. cyaneus with untreated wheat straw showed that pretreatment improved the characteristics that determine the quality of pulp. This was indicated by an increase in pulp brightness and by a decrease in the kappa number. These changes occurred without significantly affecting the viscosity, a measure of the quality of the cellulose fibres. These results support the potential application of this organism or its oxidative enzymes in biopulping.
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