Pharmaceutica Analytica Acta

CenC, a multi-domain thermo-stable GH9 processive endoglucanase from Clostridium thermocellum: Cloning, characterization and saccharification studies

International Conference and Exhibition on Advances in HPLC & Chromatography Techniques

March 14-15, 2016 London, UK

Ikram-Ul-Haq

GC University, Pakistan

Posters & Accepted Abstracts: Pharm Anal Acta

Abstract:

The growing demands of bioenergy have led to the emphasis on novel cellulases to improve efficiency of biodegradation process of plant biomass. Therefore, a thermostable cellulolytic gene (CenC) with 3,675 Bp was cloned from Clostridium thermocellum and over-expressed in Escherichia coli strain BL21 CodonPlus. It was attested that CenC belongs to glycoside hydrolase family 9 (GH9) with four binding domains, a processive endoglucanase. CenC was purified to homogeneity, producing a single band on SDSPAGE corresponding to 137.11 kDa, by purification steps of heat treatment combined with ion-exchange chromatography. Purified enzyme displayed optimal activity at pH 6.0 and 70°C. CenC had a half-life of 24 min at 74°C, was stable up to 2 h at 60°C and over a pH range of 5.5-7.5. Enzyme showed high affinity towards various substrates and processively released cellobiose from cellulosic substrates confirmed by using HPLC technique. It efficiently hydrolyzed carboxymethyl cellulose (30 U/mg), β-glucan Barley (94 U/mg); also showed activity towards p-nitrophenyl-β-D-cellobioside (18 U/mg), birchwood xylan (19 U/mg), beech wood xylan (17.5 U/mg), avicel (9 U/mg), whatman filter paper (11 U/mg) and laminarin (3.3 U/mg). CenC exhibited Km, Vmax, Kcat, VmaxKm -1 and KcatKm -1 of 7.14mM, 52.4 μmol mg-1min-1, 632.85 s-1, 7.34 min-1 and 88.63, respectively used CMC as substrate. Recombinant CenC saccharified pretreated wheat straw and bagasse to 5.12% and 7.31%, respectively at pH 7.0 and 45°C after 2 h incubation. Its thermostability, high catalytic efficiency and independence of inhibitors make CenC enzyme an appropriate candidate for industrial applications and cost-effective saccharification process.

Biography :

Email: ikmhaq@yahoo.com