Production, purification and characterization of novel beta glucosidase from newly isolated Penicillium simplicissimum H-11 in submerged fermentation

Abstract

β-Glucosidase is an important component of the cellulase complex. It not only hydrolyzes cellobiose and short-chain cellooligosaccharides to glucose, but also removes the inhibitory effect of cellobiose on the β-1, 4-endoglucanase and exoglucanase, thereby increasing the overall rate of cellulose biodegradation. β-glucosidasefrom culture supernatant of a fungus Penicillium simplicissimum was purified to homogeneity, by using ammonium sulfate fraction, Sephadex G-100 chromatography, and its properties were studied. The molecular mass of the enzyme was about 126.0 kDa, as identified by 12% SDS-PAGE. The optimum pH and temperature were 4.4 ~ 5.2 and 60 °C, respectively. The enzyme was stable in pH 5.2 ~ 6.4 and under 40 °C. Metal profile of the enzyme showed that Mn(2+) enhances its activity, while Cu(2+), Co(2+)and Fe(3+) cause obvious inhibition. The K m and V max was 14.881 mg/ml and 0.364 mg ml/min against salicin as a Substrate. This enzyme had secondary protein structure as evidenced by FTIR spectrum.

Keywords: Penicillium simplicissimum; beta-glucosidase; characterization; purification.

Table 1. Purification profile of β-glucosidase from P. simplicissimum H11

Table 2. Comparison of optimum pH and temperature of some species of genus Penicillium

Table 3. Substrate specificities of β-glucosidase

Figure 1. Sephadex G-100 elution profile of β-glucosidase

Figure 2. Purity check of β-glucosidase (a) SDS-PAGE and (b) HPLC chromatogram

Figure 3. FTIR spectrum of β-glucosidase enzyme

Figure 4. Ultraviolet absorption spectrum of β-glucosidase produced from Penicillium simplicissimum

Figure 5. Effect of temperature on β-glucosidase activity and stability

Figure 6. Effect of pH on β-glucosidase activity and stability

Figure 7. Effect of metal ions on β-glucosidase activity

Figure 8. Lineweaver-Burk plots of β-glucosidase against salicin

Figure 9. Enzymatic degradation of cellobiose by β-glucosidase enzyme produced from Penicillium simplicissimum H-11, Red line indicates standard and blue line indicates experimental.