Fortes-Dias Consuelo Latorre, Ortolani Paula Ladeira, Fernandes Carlos Alexandre H, Lobo Kelli Roberta, Amaral de Melo Lutiana, Borges Márcia Helena, Pazin Wallance Moreira, Neto Mário de Oliveira, Fernandez Roberto Morato, Fontes Marcos Roberto M
Diretoria de Pesquisa e Desenvolvimento, Fundação Ezequiel Dias (FUNED), Rua Conde Pereira Carneiro 80, CEP 30510-010, Belo Horizonte, MG, Brazil.
Diretoria de Pesquisa e Desenvolvimento, Fundação Ezequiel Dias (FUNED), Rua Conde Pereira Carneiro 80, CEP 30510-010, Belo Horizonte, MG, Brazil.
Biochim Biophys Acta. 2014 Sep;1844(9):1569-79. doi: 10.1016/j.bbapap.2014.05.001. Epub 2014 May 9.
Several snake species possess endogenous phospholipase A2 inhibitors (sbPLIs) in their blood plasma, the primary role of which is protection against an eventual presence of toxic phospholipase A2 (PLA2) from their venom glands in the circulation. These inhibitors have an oligomeric structure of, at least, three subunits and have been categorized into three classes (α, β and γ) based on their structural features. SbγPLIs have been further subdivided into two subclasses according to their hetero or homomeric nature, respectively. Despite the considerable number of sbγPLIs described, their structures and mechanisms of action are still not fully understood. In the present study, we focused on the native structure of CNF, a homomeric sbγPLI from Crotalus durissus terrificus, the South American rattlesnake. Based on the results of different biochemical and biophysical experiments, we concluded that, while the native inhibitor occurs as a mixture of oligomers, tetrameric arrangement appears to be the predominant quaternary structure. The inhibitory activity of CNF is most likely associated with this oligomeric conformation. In addition, we suggest that the CNF tetramer has a spherical shape and that tyrosinyl residues could play an important role in the oligomerization. The carbohydrate moiety, which is present in most sbγPLIs, is not essential for the inhibitory activity, oligomerization or complex formation of the CNF with the target PLA2. A minor component, comprising no more than 16% of the sample, was identified in the CNF preparations. The amino-terminal sequence of that component is similar to the B subunits of the heteromeric sbγPLIs; however, the role played by such molecule in the functionality of the CNF, if any, remains to be determined.
几种蛇类的血浆中含有内源性磷脂酶A2抑制剂（sbPLIs），其主要作用是保护机体免受循环中来自毒腺的有毒磷脂酶A2（PLA2）的侵害。这些抑制剂具有至少由三个亚基组成的寡聚结构，并根据其结构特征分为三类（α、β和γ）。SbγPLIs又根据其异源或同源性质进一步细分为两个亚类。尽管已描述的sbγPLIs数量众多，但其结构和作用机制仍未完全明确。在本研究中，我们聚焦于来自南美响尾蛇（Crotalus durissus terrificus）的同源sbγPLI——CNF的天然结构。基于不同生化和生物物理实验的结果，我们得出结论，虽然天然抑制剂以寡聚体混合物的形式存在，但四聚体排列似乎是主要的四级结构。CNF的抑制活性很可能与这种寡聚构象相关。此外，我们认为CNF四聚体呈球形，酪氨酸残基可能在寡聚化过程中发挥重要作用。大多数sbγPLIs中存在的碳水化合物部分对于CNF与靶标PLA2的抑制活性、寡聚化或复合物形成并非必需。在CNF制剂中鉴定出一种次要成分，其含量不超过样品的16%。该成分的氨基末端序列与异源sbγPLIs的B亚基相似；然而，该分子在CNF功能中所起的作用（如果有的话）仍有待确定。
