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Purification and characterisation of a β-glucosidase (cellobiase) from a mushroom Termitomyces clypeatus

1991; Elsevier BV; Volume: 1076; Issue: 2 Linguagem: Inglês

10.1016/0167-4838(91)90269-6

1878-1454

Saswati Sengupta, Anil K. Ghosh, Subhabrata Sengupta,

Enzyme-mediated dye degradation

A β-glucosidase with cellobiase activity was purified to homogeneity from the culture filtrate of the mushroom Termtomyces clypeatus. The enzyme had optimum activity at pH 5.0 and temperature 65°C and was stable up to 60° C and within pH 2–10. Among the substrates tested, p-nitrophenyl-β-d-glucopyranoside and cellobiose were hydrolysed best by the enzyme. Km and Vm values for these substrates were 0.5, 1.25 mM and 95, 91 μmol/min per mg, respectively. The enzyme had low activity towards gentiobiose, salicin and β-methyl-d-glucoside. Glucose and cellobiose inhibited the β-d-glucosidase (PNPGase) activity competitively with Ki of 1.7 and 1.9 mM, respectively. Molecular mass of the native enzyme was approximated to be 450 kDa by HPLC, whereas sodium dodecyl sulphate polyacrylamide gel electrophoresis indicated a molecular mass of 110 kDa. The high molecular weight enzyme protein was present both intracellularly and extracellularly from the very early growth phase. The enyme had a pI of 4.5 and appeared to be a glycoprotein.