Phospholipases A2 (PLA2s) are highly prevalent in Bothrops snake venom and play a crucial role in inflammatory responses and immune cell activation during envenomation. Despite their significance, the specific role of PLA2s from Bothrops mattogrossensis venom (BmV) in inflammation is not fully understood. This study sought to isolate and characterize a novel acidic PLA2 from BmV, designated BmPLA2-A, and to evaluate its effects on human umbilical vein endothelial cells (HUVECs), with a specific focus on cytotoxicity, adhesion, and detachment. BmPLA2-A was isolated through a multi-step chromatographic procedure, involving cation exchange (CM-Sepharose), hydrophobic interaction (n-butyl-Sepharose-HP), and reversed-phase (C-18) chromatography. SDS-PAGE analysis revealed a single protein band of approximately 15 kDa. The primary structure of BmPLA2-A was determined by LC-MS/MS, while its tertiary structure was modeled using AlphaFold. Enzymatic activity was verified with the synthetic substrate 4N3OBA. Molecular dynamics simulations were conducted to further investigate the catalytic mechanism of BmPLA2-A at the molecular level. In vitro assays on HUVECs revealed that BmPLA2-A neither induce cytokine release (IL-6, IL-8, IL-1β, TNF) nor affected cell viability, adhesion, or detachment. The characteristics of BmPLA2-A are consistent with those of acidic Asp-49 PLA2 enzymes, highlighting its potential involvement in the cytotoxic and inflammatory effects of the venom.
Keywords: Bothrops mattogrossensis; Endothelial cells; Phospholipase A(2); Snake venom.
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