使用亲和色谱法纯化了来自骆驼(Camelusdromedarius)红细胞的新型碳酸酐酶II(CAII),并进行了生化表征。以745.17倍的纯化和25.37%的产率获得140.88U/mg的比活性。该酶是具有较低分子量(25kDa)和较低Zn含量(每mol蛋白质0.50molZn)的单体。该酶显示出更高的最适温度(70°C)和pH(pH9.0),此外,根据热力学参数(Ea,kd,Ed,t1/2,D值,Z值,ΔH,ΔG和ΔS)。该酶被阳离子(Al3+,Ca2+,Cd2+,Co2+,Cr3+,Cu2+,Fe3+,Ni2+,Mg2+和Zn2+)以及阴离子(Br,CH3COO,ClO4,CN,F,HCO3,我,N3,NO3和SCN),一些阴离子(C6H5O73-,CO32-,SeO3和SO42-)不影响酶活性。还研究了各种化学物质对酶活性的影响。Km,Vmax,发现4-NPA的kcat和kcat/Km值为1.74mM,0.0093U/mL,分别为0.0039s-1和0.0023s-1mM-1。有了这些有趣的生化特性,骆驼CAII是严苛工业应用的有希望的候选者,特别是,一个成功的仿生二氧化碳封存过程。
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 (Ea, kd, Ed, t1/2, 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 (C6H5O73-, CO32-, SeO3‾ and SO42-) does not affect enzyme activity. Effect of various chemicals on enzyme activity was also investigated. Km, Vmax, kcat and kcat/Km 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.