Purification and biochemical characterization of a novel carbonic anhydrase II from erythrocytes of camel (Camelusdromedarius)

Abstract

A novel carbonic anhydrase II (CA II) from erythrocytes of camel (Camelus dromedarius) was purified to homogeneity using affinity chromatography and biochemically characterized. Specific activity of 140.88 U/mg was obtained with 745.17-fold purification and 25.37% yield. The enzyme was a monomer with a lower molecular weight (25 kDa) and lower Zn content (0.50 mol of Zn per mol of protein). The enzyme showed higher optimum temperature (70 °C) and pH (pH 9.0), moreover, it was stable at higher temperatures and strongly alkaline pH as judged by thermodynamic parameters (E_a, k_d, E_d, t_1/2, D-value, Z-value, ΔH, ΔG and ΔS). The enzyme was inhibited by cations (Al³⁺, Ca²⁺, Cd²⁺, Co²⁺, Cr³⁺, Cu²⁺, Fe³⁺, Ni²⁺, Mg²⁺ and Zn²⁺) as well as by anions (Br^‾, CH₃COO^‾, ClO₄^‾, CN^‾, F^‾, HCO₃^‾, I^‾, N₃^‾, NO₃^‾ and SCN^‾), some anions (C₆H₅O₇^3-, CO₃^2-, SeO₃^‾ and SO₄^2-) does not affect enzyme activity. Effect of various chemicals on enzyme activity was also investigated. K_m, V_max, k_cat and k_cat/K_m values for 4-NPA were found to be 1.74 mM, 0.0093 U/mL, 0,0039 s^-1 and 0,0023 s^-1 mM^-1, respectively. With these interesting biochemical properties, camel CA II represents promising candidate for harsh industrial applications, in particular, for a successful biomimetic CO₂ sequestration process.

Keywords: Biochemical characterization; Camel; Camelus dromedarius; Carbonic anhydrase II; Purification.