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Cellulolytic enzyme systems of Myrothecium verrucariaHash, John H. January 1957 (has links)
Crude culture filtrates of Myrothecium verrucaria. were investigated in an attempt to gain information on the following: (A) the multiplicity of the cellulolytic system, (B) the formation of glucose in the course of cellulose hydrolysis, (C) the significance of intermediate dextrins in the course of cellulose hydrolysis, (D) the properties of a β-aryl-glucosidase present in filtrates of this organism, and (E) a transglucosidase which synthesized higher saccharides from cellobiose.
In paper electrophoresis at pH 8.55 in veronal buffer of ionic strength 0.05 and a potential gradient of 10 volts/cm, concentrated culture filtrates exhibited multiple components with cellulase activity. The electrophoretic pattern for each filtrate could be reproduced; however, different filtrates gave different proportions of the components. The presence of electrophoretically distinct components with cellulase activity indicated the multiplicity of the cellulolytic system.
At pH 7.0 and lower there was no separation of the cellulase in the filtrates into the components shown at 8.55.
The β-aryl-glucosidase also exhibited several peaks on paper electrophoresis, indicating that it, too, was dependent on several proteins for its activity. With the exception of a peak of activity that remained at the origin, the mobile peaks of cellulase and β-aryl-glucosidase did not coincide, indicating that the two separate activities are not dependent on the same proteins for their respective activities. The activities remaining at the origin were also dependent on different proteins because the β-aryl-glucosidase was destroyed by heat whereas the cellulase was not.
The concentrated culture filtrates were subjected to column chromatography on alkali-swollen cellulose. The enzymes were eluted with phosphate-citrate buffers of increasing pH and ion strength. Effluent fractions were collected and assayed for their ability to hydrolyze carboxymethyl cellulose of two degrees of polymerization, insoluble cellulose dextrins, soluble cellulose dextrins, cellobiose, 6-bromo-2-naphthol-β-D-glucopyranoside, and 4-0-β-D-glucopyranosyl-D-sorbitol.
The results not o~ indicated a multienzymatic nature of cellulase but also that the components of the cellulase system were specific for a particular range of chain lengths of cellulose. Some of the fractions hydrolyzed carboxymethyl cellulose (DP 200) but did not hydrolyze the insoluble dextrins, the soluble dextrins or cellobiose; some hydrolyzed both the carboxymethyl cellulose and the insoluble dextrins but did not hydrolyze the soluble dextrins or cellobiose; and some hydrolyzed the carboxymethyl cellulose, the insoluble and soluble dextrins but not cellobiose. There were a few fractions which hydrolyzed the carboxymethyl cellulose, soluble and insoluble dextrins and cellobiose. Cellobiase was confined to a relatively few fractions and all the fractions with cellobiase activity also exhibited transglucosidase activity.
Cellulase fractions which were void of cellobiase and transglucosidase activity accumulated glucose, cellobiose, and higher saccharides when hydrolyzing insoluble cellulose dextrins. The formation of glucose in the absence of cellobiase indicated that the cellulase is capable of removing single glucose units from the end of the chain and cellobiose is not an obligatory glucose precursor. The formation of higher saccharides in the absence of transglucosidase indicated that these sugars were true intermediate dextrins, arising as the result of random cleavage of the cellulose molecule, and were not the synthetic products of a transglucosidase.
A β-aryl-glucosidase, which was not a cellobiase, chromatographed off the cellulose column. Some of the properties of this enzyme were studied. It was found to hydrolyze several β-aryl-glucosidases with varying ease. In the presence of various alcohols it transferred the glucose moiety of the glucoside to the alcohol. With methanol the product was methyl glucoside, but it was not possible to establish the configuration of the linkage that was formed.
The transglucosidase synthesized, from cellobiose, several higher saccharides which appeared to be the homologous series of cellulose dextrins through the hexasaccharide. / Ph. D.
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Phylogeny and Taxonomy of Polyblastia and allied taxa (Verrucariaceae)Savić, Sanja January 2007 (has links)
Polyblastia A. Massal. is a genus of lichenized fungi belonging to Verrucariaceae (Ascomycota). Traditional classification heavily rested on a few cardinal characters, primarily ascoma structure and spore morphology, but the delimitation of Polyblastia was problematic. Using DNA sequences from nucLSU, and RPB1 the position of the ‘Polyblastia group’, as a strongly supported clade within Verrucariaceae, was confirmed. A three marker phylogeny (including nucITS) based on a wide taxon sampling reveals strongly supported clades within the ‘Polyblastia group’. Thus Polyblastia s.str. is recognized. The phylogeny of the strongly supported Thelidium clade is not completely resolved. In a different clade, Verrucaria rupestris, the type of Verrucaria, is a sister to the Henrica melaspora/H. theleodes clade. Trimmatothele perquisita, the type of Trimmatothele, forms a strongly supported group with a Verrucaria species. The genus Sporodictyon, until recently included in Polyblastia, is resurrected. Species delimitation in Sporodictyon is investigated, and the morphological variation within and between the monophyletic groups is evaluated in an effort to reconcile morphological and molecular data. A taxonomic revision of Sporodictyon, occurring in Northern Europe and the adjacent Arctic is provided, and two new species are described. A comprehensive material is investigated, a key to the species is provided. A generic revision of the family Verrucariaceae, based on recent molecular phylogenetic analyses and previous morphological studies, is offered. Three new genera (Hydropunctaria, Parabagliettoa and Wahlenbergiella) are proposed. Several other monophyletic groups are identified for which the taxon sampling is insufficient for proposing taxonomic changes. The nomenclature of the genera in the family is revised, including the study of type materials and several lectotypifications are suggested. The new genus Atla (Verrucariaceae) is described, and three new species to science, A. alpina, A. palicei and A. praetermissa, are included. An identification key to the species and a revision of the genus is supplied.
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