Tracing the evolution of venom phospholipases A2 in Gloydius strauchii and related pitvipers: A tale of two acidic isozymes

Abstract

Two acidic Asp⁴⁹-PLA₂s with Glu⁶ substitution and a neutral Lys⁴⁹-PLA (designated Gst-K49) were cloned from G. strauchii venom glands, their full amino acid sequences were deduced. The predominant acidic PLA₂ (designated Gst-E6a) contains 124 residues and the M¹⁸W³⁰ substitutions, while the minor acidic PLA₂ (designated Gst-E6b) contains 122 residues and the V¹⁸A³⁰ substitutions. Their sequences are most similar to those of the respective orthologous PLA₂s of G. intermedius venom. Gst-E6a and Gst-E6b appear to be paralogs and possibly have different predatory targets or functions. The LC-MS/MS results indicate the presence of only three PLA₂ gene products in the crude venom, the relative expression levels were in the order of Gst-E6a ≫ Gst-E6b > Gst-K49, as confirmed by qPCR results. In contrast to other Gloydius, G. strauchii venom does not contain neurotoxic or basic anticoagulant Asp⁴⁹-PLA₂s, but Gst-K49 is the first Lys⁴⁹-PLA₂ identified in Gloydius venoms. However, its venom content is relatively low and its pI value 7.3 is much lower than those of other Lys⁴⁹-PLA₂s and. The Lys⁴⁹-PLA₂ genes appear to regress in the venom of most of Gloydius and related rattlesnake, and this evolutionary regression occurred before the dispersal of Asian pitvipers to the New World.

Keywords: Gloydius strauchii; Molecular cloning; Pitviper evolution and adaptation; Sequence alignment; Venom phospholipase A(2).