Department of Biotechnology and Bioinformatics, Jaypee University of Information Technology, Waknaghat, Solan, Himachal Pradesh, 173 234, India.
Department of Biotechnology, Himachal Pradesh University, Summer Hill, Shimla, 171 005, India.
Environ Res. 2022 Sep;212(Pt D):113538. doi: 10.1016/j.envres.2022.113538. Epub 2022 May 28.
In this study, a bacterial carbonic anhydrase (CA) was purified from Corynebacterium flavescens for the CO conversion into CaCO. The synthesized CaCO can be utilized in the papermaking industry as filler material, construction material and in steel industry. Herein, the CA was purified by using a Sephadex G-100 column chromatography having 29.00 kDa molecular mass in SDS-PAGE analysis. The purified CA showed an optimal temperature of 35 °C and pH 7.5. In addition, a kinetic study of CA using p-NPA as substrate showed V (166.66 μmoL/mL/min), K (5.12 mM), and K (80.56 sec) using Lineweaver Burk plot. The major inhibitors of CA activity were Na, K, Mn, and Al, whereas Zn and Fe slightly enhanced it. The purified CA showed a good efficacy to convert the CO into CaCO with a total conversion rate of 65.05 mg CaCO/mg of protein. In silico analysis suggested that the purified CA has conserved Zn coordinating residues such as His 111, His 113, and His 130 in the active site center. Further analysis of the CO binding site showed conserved residues such as Val 132, Val 142, Leu 196, Thr 197, and Val 205. However, a substitution has been observed where Trp 208 of its closest structural homolog T. ammonificans CA is replaced with Arg 207 of C. flavescens. The presence of a hydrophilic mutation in the CO binding hydrophobic region is a further subject of investigation.
在这项研究中，从黄色分枝杆菌中纯化了一种细菌碳酸酐酶（CA），用于将 CO 转化为 CaCO。合成的 CaCO 可在造纸工业中用作填料材料、建筑材料和钢铁工业中。在此，使用 Sephadex G-100 柱层析法纯化 CA，SDS-PAGE 分析显示其分子量为 29.00 kDa。纯化的 CA 显示出最佳温度为 35°C 和 pH 7.5。此外，使用 p-NPA 作为底物的 CA 动力学研究表明，使用 Lineweaver Burk 图，V（166.66 μmoL/mL/min）、K（5.12 mM）和 K（80.56 sec）。CA 活性的主要抑制剂是 Na、K、Mn 和 Al，而 Zn 和 Fe 则略有增强。纯化的 CA 具有将 CO 有效转化为 CaCO 的能力，总转化率为 65.05 mg CaCO/mg 蛋白质。计算机分析表明，在活性中心，纯化的 CA 具有保守的 Zn 配位残基，如 His 111、His 113 和 His 130。进一步分析 CO 结合位点显示保守残基，如 Val 132、Val 142、Leu 196、Thr 197 和 Val 205。然而，观察到一个取代，其最接近的结构同源物 T. ammonificans CA 的 Trp 208 被 C. flavescens 的 Arg 207 取代。在 CO 结合疏水区中存在亲水突变是进一步研究的主题。
