Catalytic and Thermodynamic Characterization of Endoglucanase (CMCase) from Aspergillus oryzae cmc-1
2009
Javed, Muhammad Rizwan | Rashid, Muhammad Hamid | Nadeem, Habibullah | Riaz, Muhammad | Perveen, Raheela
Monomeric extracellular endoglucanase (25 kDa) of transgenic koji (Aspergillus oryzae cmc-1) produced under submerged growth condition (7.5 U mg⁻¹ protein) was purified to homogeneity level by ammonium sulfate precipitation and various column chromatography on fast protein liquid chromatography system. Activation energy for carboxymethylcellulose (CMC) hydrolysis was 3.32 kJ mol⁻¹ at optimum temperature (55 °C), and its temperature quotient (Q ₁₀) was 1.0. The enzyme was stable over a pH range of 4.1-5.3 and gave maximum activity at pH 4.4. V max for CMC hydrolysis was 854 U mg⁻¹ protein and K m was 20 mg CMC ml⁻¹. The turnover (k cat) was 356 s⁻¹. The pK a₁ and pK a₂ of ionisable groups of active site controlling V max were 3.9 and 6.25, respectively. Thermodynamic parameters for CMC hydrolysis were as follows: ΔH* = 0.59 kJ mol⁻¹, ΔG* = 64.57 kJ mol⁻¹ and ΔS* = -195.05 J mol⁻¹ K⁻¹, respectively. Activation energy for irreversible inactivation 'E a₍d₎' of the endoglucanase was 378 kJ mol⁻¹, whereas enthalpy (ΔH*), Gibbs free energy (ΔG*) and entropy (ΔS*) of activation at 44 °C were 375.36 kJ mol⁻¹, 111.36 kJ mol⁻¹ and 833.06 J mol⁻¹ K⁻¹, respectively.
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