Characterization of thermo/halo stable cellulase produced from halophilic Virgibacillus salarius BM-02 using non-pretreated biomass

Abstract

The production of extremozymes from halophilic bacteria has increased significantly due to their stability and efficiency in catalyzing a reaction, as well as their capacity to display optimum activity at various salt concentrations. In the current study, the halophilic bacterium Virgibacillus salarius strain BM-02 could utilize many non-pretreated substrates including cellulose, corn stover, sugarcane bagasse and wheat bran as a sole carbon source. However, wheat bran was the best substrate for achieving optimum saccharification yield (90.1%). The partially purified cellulase was active and stable at a wide range of pH (5-8) with residual activities > 58%. Moreover, it was stable at 5-12% of NaCl. Metal ions have a variable impact on the activity of partially purified cellulase however, Fe⁺³ exhibited the highest increase in the cellulase activity. The enzyme exhibited a thermal stability at 40, 50 and 60 °C with half-lives of 1049.50, 168.14 and 163.5 min, respectively. The value of V_max was 22.27 U/mL while Km was 2.1 mM. The activation energy of denaturation E_d 69.81 kJ/mol, the enthalpy values (ΔH_d) were positive, and the entropy values (ΔS) were negative. Therefore, V. Salarius is recommended as a novel promising halophilic extremozyme producer and agricultural waste remover in the bio-industrial applications.

Keywords: Agricultural wastes; Cellulase; Halostable; Lignocellulosic; Thermostable; Virgibacillus sp..

Fig. 1

The correlation between the total cellular protein (µg/mL); (solid line) and the induced cellulase activity (U/mL); (dotted line) along with the time (day). Error bars represent the standard deviation (SD ±) of three replications

Fig. 2

(a) The impact of different types of agricultural wastes (20 g/L) and (b): the impact of wheat bran concentration (optimum substrate) on the cellulase activity (U/mL); (line) and the percentage of saccharification (column). Error bars represent the standard deviation (SD ±) of three replications. ^** significant difference (P ≤ 0.05)

Fig. 3

The impact of pH (a), and NaCl concentration (b) on the relative activity of partially purified cellulase. Error bars represent the standard deviation (SD ±) of three replications

Fig. 4

The Lineweaver –Burk plot to estimate Km, app and Vmax of cellulase produced by V. salaries under optimum conditions, Error bars represent the standard deviation (SD ±) of three replications

Fig. 5

The impact of temperature on the relative activity of partially purified cellulase (a), D value-plot to calculate -value (the temperature required to reduce the D-value by one logarithmic cycle) (b) and First-order Arrhenius plot for determination of activation energy of denaturation (Ed) of cellulase under the optimum enzyme conditions (c).