Background

Photocontrol of protein activity has become a helpful strategy for regulating biological pathways. Herein, a method for the precise and reversible photocontrol of oxidase activity was developed by using the conformational change of the AsLOV2 domain.

Results

The AsLOV2 domain was inserted into the nonconserved sites exposed on the surface of the AdhP protein, and the alov9 fusion was successfully screened for subsequent optical experiments under the assumption that neither of these actions affected the original activity of AdhP protein. The activity of alov9 was noticeably inhibited when the fusion was exposed to 470 nm blue light and recovered within 30 min. As a result, we could precisely and reversibly photocontrol alov9 activity through the optimization of several parameters, including cofactor concentration, light intensity, and illumination time.

Conclusions

An efficient method was developed for the photoinhibition of enzymatic activity based on the insertion of the light-sensitive AsLOV2 domain, providing new ideas for photocontrolling metabolic pathways without carriers in the future.

中文翻译：

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通过将光敏结构域插入氧化酶来对氧化酶活性进行可逆的光控制

背景

蛋白质活性的光控制已成为调节生物学途径的有用策略。本文中，通过使用AsLOV2结构域的构象变化，开发了一种精确和可逆的氧化酶活性光控制方法。

结果

将AsLOV2结构域插入暴露于AdhP蛋白表面的非保守位点，并假设这些作用均未影响AdhP蛋白的原始活性，因此成功地筛选出alov9融合蛋白用于随后的光学实验。当融合物暴露于470 nm蓝光并在30分钟内恢复时，alov9的活性受到明显抑制。结果，我们可以通过优化一些参数来精确和可逆地光控alov9活性，这些参数包括辅因子浓度，光强度和照明时间。

结论

基于光敏AsLOV2结构域的插入，开发了一种有效的酶活性光抑制方法，为将来在无载体的情况下控制代谢途径提供了新思路。