甲酸介导的酶介导的黄素腺嘌呤二核苷酸（FAD）转化为8-甲酰基FAD导致修饰的辅因子具有增强的催化性能,Biochemistry

当前位置： X-MOL 学术 › Biochemistry › 论文详情

Our official English website, www.x-mol.net, welcomes your feedback! (Note: you will need to create a separate account there.)

甲酸介导的酶介导的黄素腺嘌呤二核苷酸（FAD）转化为8-甲酰基FAD导致修饰的辅因子具有增强的催化性能
Biochemistry ( IF 2.9 ) Pub Date : 2017-07-11 00:00:00 , DOI: 10.1021/acs.biochem.7b00335
John M. Robbins _{1,

2} , Michael G. Souffrant _{3,

4,

5} , Donald Hamelberg _{3,

4,

5} , Giovanni Gadda _{3,

4,

6,

7} , Andreas S. Bommarius _{1,

2,

8}

Affiliation

黄素，包括黄素腺嘌呤二核苷酸（FAD），是基本的催化辅助因子，负责多种蛋白质的氧化还原功能。或者，在自然界中很少发现修饰的黄素类似物，因为它们的掺入通常会导致黄素蛋白失活，从而导致重要细胞途径的破坏。在这里，我们报告真菌黄酮酶甲酸氧化酶（FOX）催化非共价结合的FAD向8-甲酰基FAD的缓慢转化，这种转化导致甲酸氧化酶活性提高近10倍。尽管由于乳酸氧化酶的定点诱变研究先前已报道了结合酶的8-甲酰基FMN的存在，但FOX是野生型酶中第一个报道的8-甲酰基FAD病例。所以，使用稳态动力学，定点诱变，紫外可见，圆二色性，荧光光谱，液相色谱质谱分析和计算分析研究了甲酸氧化酶中8-甲酰基FAD辅因子的形成。令人惊讶的是，这些研究的结果不仅表明8-甲酰基FAD是自发形成的，并形成了活性形式的FOX，而且其自催化形成还取决于附近的精氨酸残基R87。因此，这项工作描述了一种新的酶辅因子，并提供了对酶介导的8α-黄素修饰的鲜为人知的机理的见解。液相色谱和质谱分析，以及计算分析。令人惊讶的是，这些研究的结果不仅表明8-甲酰基FAD是自发形成的，并形成了活性形式的FOX，而且其自催化形成还取决于附近的精氨酸残基R87。因此，这项工作描述了一种新的酶辅因子，并提供了对酶介导的8α-黄素修饰的鲜为人知的机理的见解。液相色谱和质谱分析，以及计算分析。令人惊讶的是，这些研究的结果不仅表明8-甲酰基FAD是自发形成的，并形成了活性形式的FOX，而且其自催化形成还取决于附近的精氨酸残基R87。因此，这项工作描述了一种新的酶辅因子，并提供了对酶介导的8α-黄素修饰的鲜为人知的机理的见解。

"点击查看英文标题和摘要"

Enzyme-Mediated Conversion of Flavin Adenine Dinucleotide (FAD) to 8-Formyl FAD in Formate Oxidase Results in a Modified Cofactor with Enhanced Catalytic Properties

Flavins, including flavin adenine dinucleotide (FAD), are fundamental catalytic cofactors that are responsible for the redox functionality of a diverse set of proteins. Alternatively, modified flavin analogues are rarely found in nature as their incorporation typically results in inactivation of flavoproteins, thus leading to the disruption of important cellular pathways. Here, we report that the fungal flavoenzyme formate oxidase (FOX) catalyzes the slow conversion of noncovalently bound FAD to 8-formyl FAD and that this conversion results in a nearly 10-fold increase in formate oxidase activity. Although the presence of an enzyme-bound 8-formyl FMN has been reported previously as a result of site-directed mutagenesis studies of lactate oxidase, FOX is the first reported case of 8-formyl FAD in a wild-type enzyme. Therefore, the formation of the 8-formyl FAD cofactor in formate oxidase was investigated using steady-state kinetics, site-directed mutagenesis, ultraviolet–visible, circular dichroism, and fluorescence spectroscopy, liquid chromatography with mass spectrometry, and computational analysis. Surprisingly, the results from these studies indicate not only that 8-formyl FAD forms spontaneously and results in the active form of FOX but also that its autocatalytic formation is dependent on a nearby arginine residue, R87. Thus, this work describes a new enzyme cofactor and provides insight into the little-understood mechanism of enzyme-mediated 8α-flavin modifications.

更新日期：2017-07-11

点击分享查看原文

点击收藏

阅读更多本刊新发论文本刊介绍/投稿指南