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首页> 外文期刊>Journal of Molecular Biology >In vivo generation of flavoproteins with modified cofactors.
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In vivo generation of flavoproteins with modified cofactors.

机译：在体内产生具有修饰的辅因子的黄素蛋白。

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摘要

To understand flavoprotein mechanisms and reactivity, biochemical and biophysical methods are usually employed, and differences between wild-type and mutated proteins with altered primary structures are placed under specific consideration. Alternatively, the cofactor can be modified, and modified flavoproteins can be studied accordingly. Here we present an efficient and general method for modifying the cofactor of flavoproteins in vivo. The modified cofactor is incorporated into apoprotein during protein biosynthesis in a riboflavin-auxotrophic Escherichia coli strain, which expresses a bacterial riboflavin transporter to import flavins from the medium. This system was used to introduce roseoflavin into the riboflavin-binding protein dodecin and into microbial blue-light photoreceptors of the BLUF (blue-light sensors using FAD) and LOV (light oxygen voltage) families. The modified photoreceptors showed absorption and fluorescence different from those of proteins carrying their natural cofactor or chromophores in solution, but did not show any photochemical reaction as implied by former physiological studies.
机译：为了理解黄素蛋白的机理和反应性，通常采用生化和生物物理方法，并且将具有改变的一级结构的野生型和突变型蛋白之间的差异作为特殊考虑。或者，可以修饰辅因子，并且可以相应地研究修饰的黄素蛋白。在这里，我们提出了一种在体内修饰黄素蛋白辅因子的有效且通用的方法。修饰的辅因子在核黄素营养缺陷型大肠杆菌菌株的蛋白质生物合成过程中被掺入载脂蛋白中，该菌株表达细菌核黄素转运蛋白以从培养基中导入黄素。该系统用于将玫瑰黄素引入到核黄素结合蛋白十二烷中，并引入到BLUF（使用FAD的蓝光传感器）和LOV（轻氧电压）家族的微生物蓝光感光器中。修饰的光感受器显示出与溶液中携带其天然辅因子或发色团的蛋白质不同的吸收和荧光，但未显示出以前的生理学研究所暗示的任何光化学反应。

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