GTP-BINDING PROTEIN STRUCTURE/FUNCTION STUDIES

ADP-核糖基化因子（ARF）是 多功能、多结构域的蛋白质， 在ER和高尔基体（和其他地方）的贩运，通过与 通过结合GTP激活后的脂质膜。这 促进ARF与效应物的相互作用[例如，外壳蛋白 在高尔基体衍生的I型囊泡上发现的β-COP，G蛋白 β-γ或磷脂酶D（PLD）的特定同种型]，以及 募集到膜上的蛋白质引起出芽和囊泡 阵ARF最初是通过其刺激 霍乱毒素ADP-核糖基转移酶活性。ARL蛋白是 结构与ARF非常相似，但最初认为 激活霍乱毒素，因此被命名为ARF样（ARL）。 将ARL 1定位于酿酒酵母的高尔基体后， 与囊泡运输中的作用一致， 重组rARL 1激活PLD，我们比较了ARF和 ARL的功能更加系统化。rARL 1的GTP γ S结合 而rARF 1受到以下因素的显著不同影响： 磷脂和洗涤剂，以及它们对PLD的活化， 霍乱毒素rARL 1对两者的激活是明确的，尽管 小于ARF激活。推导氨基酸比较 人ARL 1序列与其他四种人ARL序列的比较 五个人类ARF显示出最大的同一性（58%）， II类ARF。(It与人类ARF 1有57%的相同性， 早期定义的功能后果，其中一些 差异。）ARL 2和3在以下条件下未能激活PLD 对ARL 1有效。我们得出结论，ARL 1是一个连续体的一部分， ARF和ARL蛋白的结构相似， 功能，但在生理上重要的方式不同， 在许多人看来， 这是一个相当微妙的结构性差异。

ADP-ribosylation factors (ARFs) are multi-functional, multi-domain, proteins that regulate vesicular trafficking in the ER and Golgi (and elsewhere) by associating with a lipid membrane after being activated by binding GTP. This facilitates ARF interaction with effectors [e.g., the coat protein beta-COP found on type I Golgi-derived vesicles, the G-protein betagamma, or a specific isoform of phospholipase D (PLD)] and to recruit to membranes proteins that cause budding and vesicle formation. ARFs were first identified by their ability to stimulate cholera toxin ADP-ribosyltransferase activity. ARL proteins are very similar in structure to ARFs, but were initially believed not to activate cholera toxin and were, therefore, named ARF-like (ARL). After localizing ARL1 to the Golgi in Saccharomyces cerevisiae, consistent with a role in vesicular trafficking, and finding evidence of PLD activation by recombinant rARL1, we compared ARF and ARL function more systematically. GTPgammaS binding by rARL1 and rARF1 was influenced dramatically and differently by phospholipids and detergents, as was their activation of PLD and cholera toxin. Activation of both by rARL1 was unequivocal, albeit less than ARF activation. Comparison of deduced amino acid sequences of human ARL1 with those of four other human ARLs and five human ARFs revealed the greatest identity (58%) with class II ARFs. (It is 57% identical to human ARF1 and we had earlier defined functional consequences of some of those differences.) ARLs 2 and 3 failed to activate PLD under conditions effective for ARL1. We conclude that ARL1 is part of a continuum of ARF and ARL proteins that are similar in structure and probably function, but differ in ways that are physiologically important, as evidenced by the evolutionary persistence of what seem, in many instances, to be rather subtle structural differences.