时间分辨色氨酸荧光探测吡哆醛5 ' -磷酸依赖的o -乙酰丝氨酸巯基化酶的展开

Stefano Bettati , Barbara Campanini , Simona Vaccari , Andrea Mozzarelli , Giulio Schianchi , Theodore L. Hazlett , Enrico Gratton , Sara Benci
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引用次数: 9

摘要

以吡哆醛5′-磷酸为辅酶的蛋白质构成了一个大的超家族,目前分为三个功能基团和五种结构折叠类型。尽管序列和催化反应具有可变性,但它们具有相关的结构、动力学和功能特性。因此,它们构成了研究初级序列和辅酶相互作用对折叠途径、结构稳定性和酶功能的相对影响的最佳系统。o -乙酰丝氨酸巯基水解酶是一种二聚体吡哆醛5 ' -磷酸依赖酶,催化o -乙酰丝氨酸和硫合成l-半胱氨酸。本文报道的o -乙酰丝氨酸巯基水解酶展开的时间分辨荧光研究表明,辅酶稳定了蛋白质结构。全酶和载脂蛋白酶对色氨酸寿命的依赖性表明,与辅酶的相互作用在更高程度上稳定了c端结构域,而不是n端结构域。这一结果是根据辅酶带来的差异稳定和不同蛋白质区域的特殊功能作用所需的不同程度的构象灵活性之间的联系来讨论的。
本文章由计算机程序翻译,如有差异,请以英文原文为准。
Unfolding of pyridoxal 5′-phosphate-dependent O-acetylserine sulfhydrylase probed by time-resolved tryptophan fluorescence

Proteins utilizing pyridoxal 5′-phosphate as a coenzyme constitute a large superfamily and are currently classified into three functional groups and five structural fold types. Despite the variability of sequences and catalyzed reactions, they share relevant structural, dynamic and functional properties. Therefore, they constitute an optimal system to investigate the relative influence of primary sequence and coenzyme interactions on folding pathways, structural stability and enzymatic function. O-Acetylserine sulfhydrylase is a dimeric pyridoxal 5′-phosphate dependent enzyme that catalyzes the synthesis of l-cysteine from O-acetylserine and sulfide. The time-resolved fluorescence study of O-acetylserine sulfhydrylase unfolding, here reported, indicates that the coenzyme stabilizes the protein structure. The dependence on denaturant concentration of tryptophan lifetimes in the holo- and apo-enzyme demonstrates that the interactions with the coenzyme stabilize the C-terminal domain to a higher extent with respect to the N-terminal domain. This result is discussed in terms of a linkage between the differential stabilization brought about by the coenzyme and the different degrees of conformational flexibility required by the specialized functional role of distinct protein regions.

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