Band 80, Heft 1-2 - Special Issue: CAZymes Structural and functional aspects of Glycoside Hydrolases and Glycosyl Transferases; Editors: Maria-Ortiz Soto and Jürgen Seibel

The β-glucosidase enzyme was obtained from Trichoderma koningii Oudem. NRRL 54330 under optimal conditions by solid substrate fermentation (SSF) using corn cobs as substrate. The enzyme was purified by two-step procedures, ammonium sulphate precipitation and cefarose-4B- l -tyrosine-1-naphthylamine hydrophobic interaction chromatography, followed by biochemical and kinetic characterisation. The β-glucosidase was obtained from T. koningii using ground corn cob as substrate and Na 2 HPO 4 , pH 9, as humidification medium. The optimum conditions for enzyme production by SSF were 30 °C and 6 days. The purification efficiency of the obtained β-glucosidase was calculated to be 22.56-fold with a yield of 73.51 %. In the determination of β-glucosidase activity, p -nitrophenyl-β- d -glucopyranoside ( p NPG) substrate was used, and the optimum pH and temperature values at which β-glucosidase showed high activity were determined to be pH 3.0 and 75 °C. The purity of the enzyme and the presence/number of subunits were checked using two different electrophoretic methods, SDS-PAGE and NATIVE-PAGE electrophoretic methods. The K m and V max values of the purified enzyme were determined to be 0.16 mM and 2000 EU respectively. It was also found that d -(+)-glucose and δ-gluconolactone inhibitors exhibited competitive inhibition of β-glucosidase in the presence of p NPG.

Wood decomposing ascomycetes and basidiomycetes group of fungi are the most valuable microbes on the earth’s ecosystem that recycles the source of carbon; therefore, they are essential for the biorefinery industries. To understand the robustness of the enzymes and their metabolic pathways in the fungal system, label-free quantification of the total proteins was performed. The fungi showed a comparable quantity of protein abundance [ Trichoderma citrinoviride (285), Thermoascus aurantiacus (206), Ganoderma lucidum MDU-7 (102), G. lucidum (242)]. Differentially regulated proteins of ascomycetes and basidiomycetes were analyzed, and their heatmap shows upregulated and downregulated proteins [25 differentially expressed proteins in T. citrinoviride (8.62 % up-regulated and 91.37 % down-regulated) and G. lucidum (5.74 % up-regulated and 94.25 % down-regulated)] by using the normalized peptide-spectrum match (PSMs) and log2fold change. These proteins were similarly matched to the carbohydrate active enzymes family (CAZymes) like glycoside hydrolase (GH family), carbohydrate-binding module (CBM family) with auxiliary activities, and also involved in the hydrolysis of carbohydrate, lignin, xylan, polysaccharides, peptides, and oxido-reductase activity that helps in antioxidant defense mechanism. The lignocellulolytic enzymes from two different divisions of fungi and proteomics studies gave a better understanding of carbon recycling and multi-product lignocellulosic biorefinery processes.

Many disorders, including cancer and malaria, could be targeted via the pentose phosphate pathway (PPP), whose products are key in biosynthetic reactions in cells. The goal of this study was to find new PPP inhibitors. The inhibition effects of malononitrile derivatives on Glucose 6-phosphate dehydrogenase (G6PD) and 6-phosphogluconate dehydrogenase (6PGD) were analyzed through in vitro experiments. Besides, molecular docking studies were performed to predict the interactions having role in inhibition of compounds. K i constants of derivatives were found between 4.24 ± 0.46–69.63 ± 7.75 µM for G6PD and 1.91 ± 0.12–95.07 ± 11.08 µM for 6PGD. Derivatives indicated non-competitive inhibition on both enzymes except for compound 4. The findings of the molecular docking studies revealed that free-binding energy estimations agreed with in vitro data. The structure of these malononitrile derivatives may guide for drug discovery in targeting the PPP.