Thermotolerant glycosyl hydrolases-producing Bacillus aerius CMCPS1 and its saccharification efficiency on HCR-laccase (LccH)-pretreated corncob biomass
Authors: Ganesan Meena , Mathivani Vinayakamoorthy Remitha , Thankappan Sugitha , Muniraj Iniyakumar , Uthandi Sivakumar 
Published: July 2020

AbstractBackgroundThe current production of bioethanol based on lignocellulosic biomass (LCB) highly depends on thermostable enzymes and extremophiles owing to less risk of contamination. Thermophilic bacterial cellulases are preferred over fungi due to their higher growth rate, presence of complex multi-enzymes, stability, and enhanced bioconversion efficiency. Corncob, underutilized biomass, ensures energy conservation due to high lignocellulosic and more fermentable sugar content. In the present study, the thermophilic bacteriumBacillus aeriusCMCPS1, isolated from the thermal springs of Manikaran, Himachal Pradesh, India, was characterized in terms of its activity, stability, and hydrolytic capacity. A two-step process comprising: (i) a combined strategy of hydrodynamic cavitation reaction (HCR)-coupled enzymatic (LccH at 6.5 U) pretreatment for delignification and (ii) subsequent hydrolysis of pre-treated (HCR-LccH) corncob biomass (CCB) using a thermostable cocktail of CMCPS1 was adopted to validate the efficiency of the process. Some of the parameters studied include lignin reduction, cellulose increase, and saccharification efficiency.ResultAmong the five isolates obtained by in situ enrichment on various substrates,B. aeriusCMCPS1, isolated from hot springs, exhibited the maximum hydrolytic activity of 4.11. The GH activity of the CMCPS1 strain under submerged fermentation revealed maximum filter paper activity (FPA) and endoglucanase activity of 4.36 IU mL−1and 2.98 IU mL−1, respectively, at 44 h. Similarly, the isolate produced exoglucanase and β-glucosidase with an activity of 1.76 IU mL−1and 1.23 IU mL−1at 48 h, respectively. More specifically, the enzyme endo-1,4-β-dglucanase E.C.3.2.1.4 (CMCase) produced byB. aeriusCMCPS1 displayed wider stability to pH (3–9) and temperature (30–90 °C) than most fungal cellulases. Similarly, the activity of CMCase increased in the presence of organic solvents (118% at 30% acetone v/v). The partially purified CMCase from the culture supernatant of CMCPS1 registered 64% yield with twofold purification. The zymogram and SDS-PAGE analyses further confirmed the CMCase activity with an apparent molecular mass of 70 kDa. The presence of genes specific to cellulases, such as cellulose-binding domainCelB, confirmed the presence of GH family 46 and β-glucosidase activity (GH3). The multifunctional cellulases of CMCPS1 were evaluated for their saccharification efficiency on laccase (LccH, a fungal laccase fromHexagonia hirtaMSF2)-pretreated corncob in a HCR. The lignin and hemicelluloses removal efficiency of HCR-LccH was 54.1 and 6.57%, respectively, with an increase in cellulose fraction (42.25%). The saccharification efficiency of 55% was achieved with CMCPS1 multifunctional cellulases at 50 °C and pH 5.0.ConclusionThe multifunctional cellulase complex ofB. aeriusCMCPS1 is a potential biocatalyst for application in lignocellulosic biomass-based biorefineries. The saccharification ability of HCR-LccH-pretreated corncob at elevated temperatures would be an advantage for biofuel production from lignocellulosic biomass.