Fundamental features of microbial cellulose utilization are examined at successively higher levels of aggregation encompassing the structure and composition of cellulosic biomass, taxonomic diversity, cellulase enzyme systems, molecular biology of cellulase enzymes, physiology of cellulolytic microorganisms, ecological aspects of cellulase-degrading communities, and rate-limiting factors in nature. The methodological basis for studying microbial cellulose utilization is considered relative to quantification of cells and enzymes in the presence of solid substrates as well as apparatus and analysis for cellulose-grown continuous cultures. Quantitative description of cellulose hydrolysis is addressed with respect to adsorption of cellulase enzymes, rates of enzymatic hydrolysis, bioenergetics of microbial cellulose utilization, kinetics of microbial cellulose utilization, and contrasting features compared to soluble substrate kinetics. A biological perspective on processing cellulosic biomass is presented, including features of pretreated substrates and alternative process configurations. Organism development is considered for "consolidated bioprocessing" (CBP), in which the production of cellulolytic enzymes, hydrolysis of biomass, and fermentation of resulting sugars to desired products occur in one step. Two organism development strategies for CBP are examined: (i) improve product yield and tolerance in microorganisms able to utilize cellulose, or (ii) express a heterologous system for cellulose hydrolysis and utilization in microorganisms that exhibit high product yield and tolerance. A concluding discussion identifies unresolved issues pertaining to microbial cellulose utilization, suggests approaches by which such issues might be resolved, and contrasts a microbially oriented cellulose hydrolysis paradigm to the more conventional enzymatically oriented paradigm in both fundamental and applied contexts.

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Microbial cellulose utilization: fundamentals and biotechnology.

Bioavailability and bioefficacy of polyphenols in humans. I. Review of 97 bioavailability studies

http://protocol-online.org/forums/uploads/monthly_09_2010/msg-6750-042877300 1283321951.ipb

Interlaboratory testing of methods for assay of xylanase activity

Various agricultural residues, such as corn fiber, corn stover, wheat straw, rice straw, and sugarcane bagasse, contain about 20–40% hemicellulose, the second most abundant polysaccharide in nature. The conversion of hemicellulose to fuels and chemicals is problematic. In this paper, various pretreatment options as well as enzymatic saccharification of lignocellulosic biomass to fermentable sugars is reviewed. Our research dealing with the pretreatment and enzymatic saccharification of corn fiber and development of novel and improved enzymes such as endo-xylanase, β-xylosidase, and α-l-arabinofuranosidase for hemicellulose bioconversion is described. The barriers, progress, and prospects of developing an environmentally benign bioprocess for large-scale conversion of hemicellulose to fuel ethanol, xylitol, 2,3-butanediol, and other value-added fermentation products are highlighted.

Hemicellulose bioconversion

Outlook for cellulase improvement: screening and selection strategies.

By using a batch in vitro anaerobic fecal fermentation model, we have shown that the fecal microflora can rapidly deconjugate rutin, isoquercitrin, and a mixture of quercetin glucuronides High levels of beta,D-glucosidase, alpha,L-rhamnosidase, and beta,D-glucuronidase were present Rutin underwent deglycosylation, ring fission, and dehydroxylation The main metabolite, 3,4-dihydroxyphenylacetic acid, appeared rapidly (2 h) and was dehydroxylated to 3-hydroxyphenylacetic acid within 8 h The pattern of in vitro fermentation of rutin was not changed by changing the pH (60 or 69), fermentation scale (10 or 1000 mL), or donors of the inoculum Hydroxyphenylacetic acids were not methylated by colon flora in vitro The colonic microflora has enormous potential to transform flavonoids into lower molecular weight phenolics, and these might have protective biological activities in the colon The site of absorption of flavonoids and the form in which they are absorbed are critical for determining their metabolic pathway and consequent biological activities in vivo

Quercetin derivatives are deconjugated and converted to hydroxyphenylacetic acids but not methylated by human fecal flora in vitro.

Efficient secretion of two fungal cellobiohydrolases by Saccharomyces cerevisiae

Previous studies on Melanocarpus albomyces laccase have shown that this enzyme is very interesting for both basic research purposes and industrial applications. In order to obtain a reliable and efficient source for this laccase, it was produced in the filamentous fungus Trichoderma reesei. Two approaches were used: production of a non-fused laccase and a hydrophobin–laccase fusion protein. Both proteins were expressed in T. reesei under the cbh1 promoter, and significantly higher activities were obtained with the non-fused laccase in shake-flask cultures (corresponding to about 230 mg l−1). Northern blot analyses showed rather similar mRNA levels from both expression constructs. Western analysis indicated intracellular accumulation and degradation of the hydrophobin–laccase fusion protein, showing that production of the fusion was limited at the post-transcriptional level. No induction of the unfolded protein response pathway by laccase production was detected in the transformants by Northern hybridization. The most promising transformant was grown in a fermenter in batch and fed-batch modes. The highest production level obtained in the fed-batch culture was 920 mg l−1. The recombinant laccase was purified from the culture supernatant after cleaving the major contaminating protein, cellobiohydrolase I, by papain. The recombinant and wild-type laccases were compared with regard to substrate kinetics, molecular mass, pH optimum, thermostability, and processing of the N- and C-termini, and they showed very similar properties.

Expression of Melanocarpus albomyces laccase in Trichoderma reesei and characterization of the purified enzyme

Production of hemicellulolytic enzymes required in the hydrolysis of different xylans was investigated using strains of seven species of Aspergillus. Of the strains producing highest levels of xylanolytic activities, a. foetidus VTT-D-71002 was apparently non-cellulolytic and could therefore be a possible source of cellulase-free hemicellulase for applications in the pulping industry. The non-metabolizable synthetic xylobiose analogue β-methyl-D-xyloside was the best xylanase inducer of the materials tested. Batches of hemicellulase produced in laboratory scale fermentations on practical media were tested in the hydrolysis of both cellulosic and hemicellulosic substrates.

Michael Bailey

Papers

Interlaboratory testing of methods for assay of xylanase activity

Quercetin derivatives are deconjugated and converted to hydroxyphenylacetic acids but not methylated by human fecal flora in vitro.

Efficient secretion of two fungal cellobiohydrolases by Saccharomyces cerevisiae

Expression of Melanocarpus albomyces laccase in Trichoderma reesei and characterization of the purified enzyme

Production of xylanolytic enzymes by strains of Aspergillus