Sulfate Adenylation-Biochemistry & Enzymology

描述（由申请人提供）： 硫酸盐进入新陈代谢需要化学活化。唯一已知的激活硫酸盐的代谢手段是形成非常高能量的磷-硫酸酐键（？Go= -19千卡/摩尔）。这种键是活化硫酸盐（APS或PAPS）的化学标志，正是在这种高能环境中，硫酰基部分（-SO 3）迅速有利地进入其随后的代谢生物化学。活化键在ATP硫酸化酶催化的转移反应中形成，其中ATP的腺苷酰部分（AMP~）转移为硫酸根。 在哺乳动物中，硫酰基转移到蛋白质和小分子代谢物调节各种各样的代谢过程，包括神经肽和类固醇激素作用，生长因子识别和淋巴细胞循环。该提案概述了结构为基础的机制调查，旨在解决核心问题的功能和进化的哺乳动物类ATP磺酰化酶。 细菌ATP硫酰化酶含有一个GT3亚基（在本实验室发现），它是延伸因子Tu的进化后代。 该亚基由于GTP水解而经历的构象变化加速了腺苷酰转移酶亚基的周转，并且耦合了GTP水解和APS合成的化学势。 我们最近发现ATP硫酸化酶与半胱氨酸生物合成途径中的另一种酶（O-乙酰-L-丝氨酸巯基解酶）形成复合物，并且它们的相互作用产生“新”催化功能-ATP的水解。 这些酶组织成代谢泵，每次冲程将一个APS分子输送到通路中。 该基金将探讨该泵的机制。 与一个尚未表征和新的ATP硫酸化酶从结核分枝杆菌的工作，我们的初步数据扩展这些发现，包括五个酶的途径。 我们将定义和研究半胱氨酸代谢子，目的是了解从这一途径的自组织中出现的分层功能。

DESCRIPTION (provided by applicant): The entry of sulfate into metabolism requires that it be chemically activated. The only known metabolic means of activating sulfate is the formation of the very high-energy phosphoric-sulfuric acid anhydride bond (?Go= -19 kcal/mole). This bond is the chemical hallmark of activated sulfate (APS or PAPS), and it is from this high-energy environment that the sulfuryl-moiety (-SO3) passes quickly and favorably into its subsequent metabolic biochemistry. The activated bond is formed in a transfer reaction, catalyzed by ATP sulfurylase, in which the adenylyl-moiety (AMP~) of ATP is transferred to sulfate. In mammals, sulfuryl-group transfer to proteins and small molecule metabolites regulates a wide-variety of metabolic processes including neuropeptide- and steroid-hormone action, growth-factor recognition, and lymph cell circulation. This proposal outlines structurally-based mechanistic inquires designed to address central issues regarding the function and evolution of the mammalian class of ATP sulfurylases. Bacterial ATP sulfurylases harbor a GTPase subunit (discovered in this laboratory) that is an evolutionary descendant of elongation factor Tu. The conformational changes that this subunit undergoes as a consequence of GTP hydrolysis accelerate turnover of the adenylyl-transferase subunit, and couple the chemical potentials of GTP hydrolysis and APS synthesis. We have recently discovered that ATP sulfurylase forms a complex with another enzyme in the cysteine biosynthetic pathway (O-acetly-l-serine sulfhydrylase), and that their interactions produce "new" catalytic function - the hydrolysis of ATP. These enzymes organize into a metabolic pump, each stroke of which delivers one molecule of APS into the pathway. The mechanism of the pump will be explored in this grant. Working with an as yet uncharacterized and novel ATP sulfurylase from Mycobacterium tuberculosis, our preliminary data extends these finding to include five of the seven enzymes in the pathway. We will define and study the cysteine metabolon with the goal of understanding the hierarchical functions that emerge from the self-organization of this pathway.