Profiling and production of hemicellulases by thermophilic fungus Malbranchea flava and the role of xylanases in improved bioconversion of pretreated lignocellulosics to ethanol.

Manju Sharma,Bhupinder Singh Chadha,Chhavi Mahajan,Manpreet S Bhatti

doi:10.1007/s13205-015-0325-2

Abstract

This study reports thermophilic fungus Malbranchea flava as a potent source of xylanase and xylan-debranching accessory enzymes. M. flava produced high levels of xylanase on sorghum straw containing solidified culture medium. The optimization of culture conditions for production of hemicellulases was carried out using one factor at a time approach and Box–Behnken design of experiments with casein (%), inoculum age (h) and inoculum level (ml) as process variables and xylanase, β-xylosidase, acetyl esterases and arabinofuranosidase as response variables. The results showed that casein concentration between 3.0 and 3.5 %, inoculum age (56–60 h) and inoculum level (2–2.5 ml) resulted in production of 16,978, 10.0, 67.7 and 3.8 (U/gds) of xylanase, β-xylosidase, acetyl esterase and α-l-arabinofuranosidase, respectively. Under optimized conditions M. flava produced eight functionally diverse xylanases with distinct substrate specificity against different xylan types. The peptide mass fingerprinting of 2-D gel electrophoresis resolved proteins indicated to the presence of cellobiose dehydrogenase and glycosyl hydrolases suggesting the potential of this strain in oxidative and classical cellulase-mediated hydrolysis of lignocellulosics. Addition of xylanase (300 U/g substrate) during saccharification (at 15 % substrate loading) of different pretreated (acid/alkali) substrates (cotton stalks, wheat straw, rice straw, carrot grass) by commercial cellulase (NS28066) resulted in 9–36 % increase in saccharification and subsequent fermentation to ethanol when compared to experiment with commercial enzyme only. High ethanol level 46 (g/l) was achieved with acid pretreated cotton stalk when M. flava xylanase was supplemented as compared to 39 (g/l) with xylanase without xylanase addition.Electronic supplementary materialThe online version of this article (doi:10.1007/s13205-015-0325-2) contains supplementary material, which is available to authorized users.

Highlights

Hemicellulose is the second most abundant biopolymer in plant cell wall after cellulose which exists as O-acetyl-4O-methylglucuronoxylan in hardwoods and as arabino-4O-methylglucuronoxylan in softwoods, while xylan in grasses and annual plants are typically arabinoxylans consisting of a b-1,4-linked backbone of D-xylopyranosyl residues to which a-L-arabinofuranosyl residues are linked at C-3 and C-2 (Scheller and Ulvskov 2010)
This study reports thermophilic fungus Malbranchea flava as a potent source of xylanase and xylandebranching accessory enzymes
Malbranchea flava isolated from composting soil was found to utilize different complex carbon sources mainly derived from agro-residues for xylanase production

Summary

Introduction

Hemicellulose is the second most abundant biopolymer in plant cell wall after cellulose which exists as O-acetyl-4O-methylglucuronoxylan in hardwoods and as arabino-4O-methylglucuronoxylan in softwoods, while xylan in grasses and annual plants are typically arabinoxylans consisting of a b-1,4-linked backbone of D-xylopyranosyl residues to which a-L-arabinofuranosyl (araf) residues are linked at C-3 and C-2 (Scheller and Ulvskov 2010). Xylanases have received a great deal of research attention because of their biotechnological potential in food, feed, and pre-bleaching of pulps in paper industries. Each of these applications do require xylanases with distinct physico-chemical properties in terms of their mode of action, substrate specificity, pH, temperature optima, etc. Recent reports on xylanases (derived from recombinant T. reesei MultifectÒ) suggest important role of these xylanases in enhancing the capability of the cellulases for hydrolysis of pretreated substrates (Hu et al 2013). Owing to increasing biotechnological importance of thermostable xylanases, there is a need to identify novel and catalytic efficient sources of xylanases for their role in improving these processes. The role of M. flava xylanases in enhancing saccharification of differently pretreated substrates for subsequent fermentation to ethanol was examined

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Results

Conclusion