Salt-tolerant mechanism of marine Aspergillus niger cellulase cocktail and improvement of its activity

The cellulase cocktail produced by marine Aspergillus niger exhibits a property of salt-tolerance,which is of great potential in cellulose degradation in high salt environment.In order to explain the mechanism on the salttolerance of the cellulase cocktail produced by marine A.niger,six cellulase components(AnCel6,AnCel7A,AnCel7B,AnEGL,AnBGL1 and AnBGL2)were obtained by directed expression.Studies on their enzymatic properties revealed that oneβ-glucosidase(AnBGL2)and one endoglucanase(AnEGL)exhibited an outstanding salttolerant property,and one cellobiohydrolase(AnCel7B)exhibited a certain salt-tolerant property.Subsequent study revealed that the salt-tolerant An EGL and AnCel7B endowed the cellulase cocktail with stronger salttolerant property,while the salt-tolerant An BGL2 had no positive effect.Moreover,after overexpression of AnCel6,AnCel7A,AnCel7B and AnEGL,the activity of cellulase cocktail increased by 80%,70%,63%and 68%,respectively.However,the activity of cellulase cocktail was not improved after overexpression of AnBGL1 and AnBGL2.After mixed-strain fermentation with cellobiohydrolase recombinants(cel6 a,cel7a and cel7b recombinants)and endoglucanase recombinant(egl recombinant),the the activity of cellulase cocktail increased by 114%,102%and91%,respectively.

A boronate-modified renewable nanointerface for ultrasensitive electrochemical assay of cellulase activity

The saccharification of cellulosic biomass to produce biofuels and chemicals is one of the most promising industries for gree n-power production and sustainable development.Cellulase is the core component in the saccharification process.Simple and efficient assay method to determine cellulase activity in saccharification is thus highly required.In this work,a boronate-affinity surface based renewable and ultrasensitive electrochemical sensor for cellulase activity determination has been fabricated.Through bo ronate-sugar interaction,celluloses are attached to the electrode surface,forming the cellulose na nonetwork at the sensing interface.Cellulase degradation can lead to the variation of electrochemical impedance.Thus,electrochemical impedance signal can reflect the cellulase activity.Importantly,via fully utilizing the boronate-affinity chemistry that enables reversible fabrication of cellulose nanonetwork,a renewable sensing surface has been firstly constructed for cellulase activity assay.Thanks to interfacial diffusion process of electrochemical sensor,the product inhibitory effect in the cellulase activity assays can be circumvented.The proposed electrochemical sensor is ultrasensitive for label-free cellulase activity detection with a very simple fabrication process,showing great potential for activity screen of new enzymes in saccharification conversion.

Two-step fermentation process and the corresponding study on its enzymology characteristics

Strain HIT-3,which has good performances in cellulose degradation,was isolated from the campus soil. Based on the identification of conservative sequences 16S rDNA and the analysis of physiological-biochemical characteristics,HIT-3 was identified as Achromobacter xylosoxidans. Denitrificans. A two-step fermentation process was conducted by adopting compound-bioflocculant-producing flora constructed by cellulose-degrading bacterium HIT-3 and flocculating bacterium F2. The cellulose degradation metabolites of HIT-3 was taken as substrates by flocculating bacterium F2,by which excellent compound bio-flocculant was obtained. In addition,the enzymology characteristics of HIT-3 were investigated when cultured in cellulose media,which utilized CMC-Na as its sole carbon. The results show that HIT-3 achieves a climax of 67.6 U/mL of enzyme production after incubation for 6 d,and the organic carbons produced are sufficient as the substrates required by the fermentation of flocculating bacterium F2(flocculating efficiency of 85.6%),which makes it feasible to reuse bioenergy.

Screening and Identification of Efficient Cellulose-degrading Strain and Its Preliminary Application in Sheep Dung Compost

The paper was to solve the problems of incomplete fermentation and more cellulose impurities in finished products caused by high cellulose content in sheep dung and accessory substances in composting production.[Method]Efficient cellulose-degrading strains were screened from different animal dung samples and three widely used commercial microbial composting strains(RW,BDM and JBB).The target strains were comprehensively screened by Congo red staining,degradation test of filter strips and cellulase activity test,and the strains screened were identified by physiological and biochemical tests and molecular tests.The strain was used to produce bioaugmentation strain,which was compared with composting fermentation of commercial strains.The temperature of composting process and the physical and chemical properties of finished products were tested to determine whether it met the national standards.[Result]A strain(M2)with strong cellulose-degrading ability from horse dung was screened,and its cellulase activity was(126.06±0.36)U/mL,higher than that of cellulose strain with the highest enzyme activity in commercial strains.Through morphological,physiological and biochemical identification and 16S rDNA sequence analysis,strain M2 was identified as Sphingomonas paucimobilis belonging to Sphingomonas sp.Compared with commercial strains,the bioaugmentation strain effectively improved the cellulose-degrading ability of strains after composting fermentation.The products were uniform in color,without obvious cellulose impurities,and the total nutrients were improved to a certain extent.[Conclusion]The strain M2 screened can be used for the development of specific strain for sheep dung composting.

Screening and Characterization of the High-Cellulase-Producing Strain Aspergillus glaucus XC9

Cellulose is a kind of renewable resource that is abundant in nature.It can be degraded by microorganisms such as mildew.A mildew strain with high cellulase activity was isolated from mildewy maize cob and classified as Aspergillus glaucus XC9 by morphological and 18S rRNA gene sequence analyses.We studied the effects of nitrogen source,initial pH,temperature,incubation time,medium composition,and surfactants on cellulase production.Maximal activities of carboxymethylcellulase(6,812 U/g dry koji)and filter paperase(172 U/g dry koji)were obtained in conditions as follows:initial pH,5.5–6.0;temperature,30℃;cultivation period,3–4 days;inoculum ratio,6%(vol/vol);sugarcane bagasse/wheat bran ratio,4:6.When bagasse was used as substrate and mixed with wet koji at a 1:1(wt/wt)ratio,the yield of reducing sugars was 36.4%.The corresponding conversion rate of cellulose to reducing sugars went as high as 81.9%.The results suggest that A.glaucus XC9 is a preferred candidate for cellulase production.