Lee W H, da Silva Giotto M T, Marangoni S, Toyama M H, Polikarpov I, Garratt R C
Laboratório Nacional de Luz Síncrotron, Caixa Postal 6192, 13083-970 Campinas, São Paulo, Brazil.
Biochemistry. 2001 Jan 9;40(1):28-36. doi: 10.1021/bi0010470.
Asp49 plays a fundamental role in supporting catalysis by phospholipases A2 by coordinating the calcium ion which aids in the stabilization of the tetrahedral intermediate. In several myotoxins from the venoms of Viperidae snakes, this aspartic acid is substituted by lysine. The loss of calcium binding capacity by these mutants has become regarded as the standard explanation for their greatly reduced or nonexistent phospholipolytic activity. Here we describe the crystal structure of one such Lys49 PLA2, piratoxin-II, in which a fatty acid molecule is observed within the substrate channel. This suggests that such toxins may be active enzymes in which catalysis is interrupted at the stage of substrate release. Comparison of the present structure with other PLA2s, both active and inactive, identifies Lys122 as one of the likely causes of the increased affinity for fatty acid in Lys49 enzymes. Its interaction with the mainchain carbonyl of Cys29 is expected to lead to hyperpolarization of the peptide bond between residues 29 and 30 leading to an increased affinity for the fatty acid headgroup. This strongly bound fatty acid may serve as an anchor to secure the toxin within the membrane thus facilitating its pathological effects.
天冬氨酸49在支持磷脂酶A2的催化过程中起着重要作用，它通过配位钙离子来帮助稳定四面体中间体。在蝰蛇科毒蛇毒液中的几种肌毒素中，这种天冬氨酸被赖氨酸取代。这些突变体失去钙离子结合能力，已被视为其磷脂水解活性大大降低或不存在的标准解释。在此，我们描述了一种这样的赖氨酸49型磷脂酶A2——pir毒素-II的晶体结构，其中在底物通道内观察到一个脂肪酸分子。这表明此类毒素可能是活性酶，只是在底物释放阶段催化作用被中断。将目前的结构与其他活性和非活性磷脂酶A2进行比较，确定赖氨酸122是赖氨酸49型酶中脂肪酸亲和力增加的可能原因之一。预计它与半胱氨酸29的主链羰基相互作用会导致29和30位残基之间的肽键超极化，从而增加对脂肪酸头部基团的亲和力。这种紧密结合的脂肪酸可能作为一个锚，将毒素固定在膜内，从而促进其病理作用。
