Anoxic Enzymatic Conversion of Acetylene

乙炔的缺氧酶促转化

• 批准号: 210678598
• 负责人: Professor Dr. Oliver Einsle
• 金额: --
• 依托单位: Institut für Biochemie
• 依托单位国家: 德国
• 项目类别: Priority Programmes
• 财政年份: 2011
• 资助国家: 德国
• 起止时间: 2010-12-31 至 2014-12-31
• 项目状态: 已结题
• 来源: https://gepris.dfg.de/gepris/projekt/210678598?language=en
• 关键词: Anoxic; Enzymatic; Conversion; Acetylene

The gas acetylene (ethyne) undergoes two known enzymatic conversions in nature. Nitrogenase, the enzyme required for the reductive fixation of dinitrogen into bioavailable ammonium, reduces acetylene in a two-electron step to ethylene (ethene). In contrast, the tungsten / [4Fe:4S] enzyme acetylene hydratase catalyzes the hydratation of the gas to yield acetaldehyde. The proteins are well characterized structurally and functionally, but the mode and exact site of acetylene binding and the mechanism of its conversion remain under heavy debate for both cases. Clearly, the mechanisms of acetylene conversion follow entirely different routes: while nitrogenase most likely binds acetylene (and other substrates) to the [Mo:7Fe:9S:X]:homocitrate FeMo cofactor, from which electrons of a low redox potential get transferred directly to the substrate, the tungstoprotein acetylene hydratase has a distinct, binding pocket pre-formed by a ring of hydrophobic amino acids. Here the reaction may not involve the formation of a metal-carbon bond, but rather the activation of a water molecule coordinated to tungsten that subsequently attacks the triple bond of the alkyne. We will establish and optimize both native and recombinant production of acetylene hydratase in our laboratory, create point-specific mutants that will be analyzed for enzymatic activity and obtain high-resolution structures of wild type and variant proteins pressurized with acetylene or other candidate ligands. Nitrogenase will be isolated from Azotobacter vinelandii and assayed for acetylene reduction activity. Acetylene, and in particular the strong ligand CO will be used in pressurization experiments to obtain atomic resolution data from which we expect to gain essential mechanistic clues.