Reliable simultaneous zymographic method of characterization of cellulolytic enzymes from fungal cellulase complex

Dojnov, Biljana; Grujić, Marica; Vujčić, Zoran

doi:10.1002/elps.201400541

Cited by 6 publications

(3 citation statements)

References 19 publications

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“…This has previously been reported in both Trichoderma sp. and in Bjerkandera adusta where different cellulolytic profiles are produced when the fungi are grown on various natural substrates [105,117]. …”

Section: Resultsmentioning

confidence: 99%

Characterization of lignocellulolytic activities from fungi isolated from the deep-sea sponge Stelletta normani

et al. 2017

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Extreme habitats have usually been regarded as a source of microorganisms that possess robust proteins that help enable them to survive in such harsh conditions. The deep sea can be considered an extreme habitat due to low temperatures (<5°C) and high pressure, however marine sponges survive in these habitats. While bacteria derived from deep-sea marine sponges have been studied, much less information is available on fungal biodiversity associated with these sponges. Following screening of fourteen fungi isolated from the deep-sea sponge Stelletta normani sampled at a depth of 751 metres, three halotolerant strains (TS2, TS11 and TS12) were identified which displayed high CMCase and xylanase activities. Molecular based taxonomic approaches identified these strains as Cadophora sp. TS2, Emericellopsis sp. TS11 and Pseudogymnoascus sp. TS 12. These three fungi displayed psychrotolerance and halotolerant growth on CMC and xylan as sole carbon sources, with optimal growth rates at 20°C. They produced CMCase and xylanase activities, which displayed optimal temperature and pH values of between 50–70°C and pH 5–8 respectively, together with good thermostability and halotolerance. In solid-state fermentations TS2, TS11 and TS12 produced CMCases, xylanases and peroxidase/phenol oxidases when grown on corn stover and wheat straw. This is the first time that CMCase, xylanase and peroxidase/phenol oxidase activities have been reported in these three fungal genera isolated from a marine sponge. Given the biochemical characteristics of these ligninolytic enzymes it is likely that they may prove useful in future biomass conversion strategies involving lignocellulosic materials.

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Section: Resultsmentioning

confidence: 99%

Characterization of lignocellulolytic activities from fungi isolated from the deep-sea sponge Stelletta normani

et al. 2017

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“…The comparison of cellulase-producing strains is very important for industrial characterisation of an entire secretome, as well as for the more fundamental study of cellulose expression. Each cellulase producer has its own cellulase profile that is significant from an industrial point of view (Dojnov et al 2015).…”

Section: Discussionmentioning

confidence: 99%

Screening of new secretory cellulases from different supernatants of white rot fungi from Misiones, Argentina

et al. 2016

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Cellulases hydrolyse the cellulose chain into single sugars efficiently. These sugars can be fermented in the bioethanol process, a source of renewable energy. Misiones rainforest is one of the most biodiverse systems on the planet subtropical ecoregions, which is the most probable site to find new fungal strains with potential for degrading cellulose through cellulases. The aim of this work was to find an efficient cellulolytic microorganism through the exploration of native white rot fungi from Misiones. From the qualitative screening 11 fungal strains were selected. The quantitative analysis revealed that the isolated LBM 033 was the best cellulases producer, reaching 57, 226 and 387 U/l of cellobiohydrolase, β-glucosidase and endoglucanase activity, respectively. The zymograms showed that the molecular mass of most of the endoglucanases ranged from 69 to 88 kDa and the molecular mass of most of the cellobiohydrolases was 45 kDa. The search of new cellulases of secretory organisms should lead to an efficient degradation of cellulosic materials, and thus facilitating potential applications in the production of bioenergy from lignocellulosic biomass.

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“…The zymogram method was developed for studying hydrolytic enzymes on the basis of substrate degradation. Since the first zymogram method was described for the detection of endocellulase and endoxylanase activities , different variations have been developed , including protein separation under non‐denaturing , semi‐denaturing , denaturing conditions, and protein separation based on isoelectric focusing have been reported. Nevertheless, there are few zymogram methods for protein separation under denaturing conditions, molecular weight (MW) estimation, and detection of cellulase or xylanase activity in the same gel .…”

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confidence: 99%

One‐step zymogram method for the simultaneous detection of cellulase/xylanase activity and molecular weight estimation of the enzyme

et al. 2016

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Here, we describe a zymographic method for the simultaneous detection of enzymatic activity and molecular weight (MW) estimation, following a single electrophoresis step. This involved separating cellulase and xylanase activities from bacteria and fungi, obtained from different sources, such as commercial extracts, crude extract and purified proteins, under denaturing conditions, by 10% polyacrylamide gel electrophoresis, using polyacrylamide gels copolymerized with 1% (w/v) carboxymethylcellulose or beechwood xylan as substrates. Then, enzymes were refolded by treatment with 2.5% Triton X-100 in an appropriate buffer for each enzymatic activity, and visualized by Coomassie blue staining for MW estimation. Finally, Congo red staining revealed bio-active cellulase and xylanase bands after electrophoretic separation of the proteins in the preparations. This method may provide a useful additional tool for screening of particular cellulase and xylanase producers, identification and MW estimation of polypeptides that manifest these activities, and for monitoring and control of fungal and bacterial cellulase and xylanase production.

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Reliable simultaneous zymographic method of characterization of cellulolytic enzymes from fungal cellulase complex

Cited by 6 publications

References 19 publications

Characterization of lignocellulolytic activities from fungi isolated from the deep-sea sponge Stelletta normani

Characterization of lignocellulolytic activities from fungi isolated from the deep-sea sponge Stelletta normani

Screening of new secretory cellulases from different supernatants of white rot fungi from Misiones, Argentina

One‐step zymogram method for the simultaneous detection of cellulase/xylanase activity and molecular weight estimation of the enzyme

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