Evaluation of Glycosyl Hydrolases in the Secretome of Aspergillus fumigatus and Saccharification of Alkali-Treated Rice Straw

Abstract

A thermotolerant Aspergillus fumigatus strain isolated from composting pile of mixed industrial waste was found to produce a spectrum of cellulase and hemicellulases when cultured on rice straw solidified substrate. The two-dimensional electrophoresis (2DE) resolved the secretome into 57 distinct protein spots. The zymograms developed against 2DE gels identified the presence of three β-glucosidases and five CBHI/EGI isoforms in the secretome. The peptide mass fingerprinting of 17 protein spots by liquid chromatography mass spectrometry characterized the secretome into different glycosyl hydrolase families. The enzyme cocktail produced by A. fumigatus was capable of efficient hydrolysis of alkali pretreated rice straw (at 7% and 10% w/v) resulting in 95% and 91% saccharification, respectively.

Penicillium janthinellum: a Source of Efficient and High Levels of β-Glucosidase

“…2a). A similar approach has also been previously reported for localizing active β-glucosidase and other glycosyl hydrolases [7,17]. Twenty-four distinct protein spots from 2DE gel were excised (Fig.…”

“…As many as 14 proteins were identified as hypothetical proteins. Peptide mass fingerprinting of the proteins resolved by 2DE has proven as a powerful tool for secretome analysis and characterizing distinct glycosyl hydrolases produced by important [7,18,19]. This is the first report on the proteome analysis of P. janthinellum.…”

“…After equilibration, the IPG strips were blotted to remove excess equilibration buffer and then placed onto 12 % SDS polyacrylamide gels and overlaid with 0.5 % low-melting agarose solution containing traces of BPB. The second dimension separation was carried out at constant 25 mA current using the Hoefer mini VE system (GE Healthcare Biosciences) [7]. The resolved protein spots were developed using the silver staining method.…”

Newly isolated Penicillium oxalicum A592-4B secretes enzymes that degrade milled rice straw with high efficiency

“…For example, conversion rates of 80-90% can be achieved by pretreatment with dilute sulfuric acid, 23) dilute NaOH, 24,25) and AFEX. 26) This is because chemical treatments decrease lignin and hemicellulose contents and cause a swelling effect of cellulose in addition to the effects of milling treatments such as increased surface area and reduced cellulose crystallinity.…”

α-L-ARABINOFURANOSIDASE FROM AN EFFICIENT HEMICELLULOLYTIC FUNGUS Penicillium janthinellum CAPABLE OF HYDROLYZING WHEAT AND RYE ARABINOXYLAN TO ARABINOSE

“…One unit of xylanase activity was expressed as the amount of enzyme required to release 1 μmole of xylose per min under the assay conditions. The substrates, pNP-β-D-xylopyranoside, pNP-α-Larabinofuranoside, (3 mM) prepared in sodium acetate buffer (50 mM, pH 5.0) were used to assay β-xylosidase, α-L-arabinofuranosidase (AFase), respectively using microtitre plate based method (Sharma et al, 2011). A reaction mixture (100µl) containing 50µl of sodium acetate buffer (50mM, pH 5.0), 25µl of suitably diluted enzyme, 25 µl of substrate (3mM) was incubated at 50 o C for 30 min.…”