School of Chemical Science and Engineering, Tongji University, Shanghai, 200092, China.
Shanghai Key Laboratory of Chemical Assessment and Sustainability, Tongji University, Shanghai, 200092, China.
Biodegradation. 2024 Oct 29;36(1):2. doi: 10.1007/s10532-024-10102-4.
Manganese is an essential trace element for humans, animals, and plants, but excessive amounts of manganese can cause serious harm to organisms. The biological manganese oxidation process mainly oxidizes Mn(II) through the secretion of unique manganese oxidase by manganese-oxidizing bacteria. The T1 Cu site of multicopper oxidase is the main site for substrate oxidation, and its role is to transfer electrons to TNC, where dioxygen reduction occurs. In this study, methionine (Met) No. 444 interacting with the T1Cu-coordinating amino acid in the multicopper oxidase CopA from Brevibacillus panacihumi MK-8 was mutated to phenylalanine (Phe) and leucine (Leu) by the enzyme. Based on the analysis of enzymatic properties and the structural model, the mutant protein M444F with 4.58 times the catalytic efficiency of the original protein CopA and the mutant protein M444L with 1.67 times the catalytic efficiency of the original protein CopA were obtained. The study showed that the manganese removal rate of the manganese-oxidizing engineered bacterium Rosetta-pET-copA cultured for 7 days was 88.87%, which was 10.77% higher than that of the original engineered bacterium. Overall, this study provides a possibility for the application of genetic engineering in the field of biological manganese removal.
锰是人类、动物和植物必需的微量元素，但过量的锰会对生物体造成严重的伤害。生物锰氧化过程主要通过锰氧化菌分泌的独特锰氧化酶将 Mn(II)氧化。多铜氧化酶的 T1Cu 位是底物氧化的主要位点，其作用是将电子转移到 TNC，在那里发生氧气还原。在这项研究中，通过酶将 Brevibacillus panacihumi MK-8 中的多铜氧化酶 CopA 中与 T1Cu 配位氨基酸相互作用的蛋氨酸（Met）444 突变为苯丙氨酸（Phe）和亮氨酸（Leu）。基于酶学性质和结构模型的分析，获得了突变蛋白 M444F，其催化效率是原始蛋白 CopA 的 4.58 倍，突变蛋白 M444L 的催化效率是原始蛋白 CopA 的 1.67 倍。研究表明，培养 7 天的锰氧化工程菌 Rosetta-pET-copA 的锰去除率为 88.87%，比原始工程菌提高了 10.77%。总体而言，这项研究为遗传工程在生物除锰领域的应用提供了可能性。
