权益分类	功能权益	普通用户	{{item.name}}会员
{{category.name}}	{{benefitItem.name}}

Heterologous Production and Characterization of Multiheme Cytochrome c Enzymes

多血红素细胞色素 c 酶的异源生产和表征

基本信息

批准号：
186439814
负责人：
Professor Dr. Oliver Einsle
金额：
--
依托单位：
Arbeitsgruppe Microbial Energy Conversion and Biotechnology
依托单位国家：
德国
项目类别：
Research Grants
财政年份：
2010
资助国家：
德国
起止时间：
2009-12-31 至 2015-12-31
项目状态：
已结题

来源：
https://gepris.dfg.de/gepris/projekt/186439814?language=en
关键词：
Heterologous Production Characterization Multiheme Cytochrome

项目摘要

Multiheme Cytochromes c are a diverse family of electron carriers and redox enzymes that play a central role in several metabolic pathways of the biogeochemical nitrogen cycle. Their heme cofactors pack into conserved interaction motifs that allow for strong electronic cou-pling and efficient electron transfer, but thus also complicate spectroscopic investigations. The enzymes nitrite reductase, hydroxylamine oxidoreductase and hydrazine oxidoreductase share a highly conserved heme group arrangement and a similar architecture of the respec-tive active site, yet they catalyze very different chemistry. Knowledge about these systems is on very different levels, but to date no concise, comparative characterization of their sub-strate and product ranges has been carried out. We thus propose to combine the cyto-chromes c already under study in our laboratories with novel enzymes for which production protocols will be established and to elevate our knowledge on these systems to a common level that includes (i) the establishment of gene expression protocols for recombinant cyto-chrome c production, (ii) functional assays to probe the substrate and product range, (iii) structural characterization by X-ray crystallography and (iv) functional modification by site-directed mutagenesis. Single crystal UV/vis and EPR spectroscopy will be used to identify individual heme groups and to study intramolecular electron transfer events.

多血红素细胞色素c是一个多样化的电子载体和氧化还原酶家族，在生物地球化学氮循环的几种代谢途径中发挥核心作用。它们的血红素辅因子包装成保守的相互作用基序，允许强电子耦合和有效的电子转移，但因此也使光谱研究复杂化。亚硝酸盐还原酶、羟胺氧化还原酶和肼氧化还原酶具有高度保守的血红素基团排列和各自活性位点的相似结构，但它们催化的化学性质却截然不同。关于这些系统的知识处于非常不同的水平，但迄今为止，还没有对它们的基材和产品范围进行简明、比较的描述。因此，我们建议将我们实验室已经在研究的细胞色素c与将建立生产方案的新型酶结合起来，并将我们对这些系统的知识提升到一个共同的水平，包括(i)建立重组细胞色素c生产的基因表达方案，（ii）功能测定以探测底物和产品范围。（iii）通过x射线晶体学进行结构表征；（iv）通过定点诱变进行功能修饰。单晶紫外/可见和EPR光谱将用于识别单个血红素基团和研究分子内电子转移事件。