Venom Proteomics of Trimeresurus gracilis, a Taiwan-Endemic Pitviper, and Comparison of Its Venom Proteome and VEGF and CRISP Sequences with Those of the Most Related Species

Toxins (Basel). 2023 Jun 22;15(7):408. doi: 10.3390/toxins15070408.

Abstract

Trimeresurus gracilis is an endemic alpine pitviper in Taiwan with controversial phylogeny, and its venom proteome remains unknown. In this study, we conducted a proteomic analysis of T. gracilis venom using high-performance liquid chromatography-tandem mass spectrometry and identified 155 toxin proteoforms that belong to 13 viperid venom toxin families. By searching the sequences of trypsin-digested peptides of the separated HPLC fractions against the NCBI database, T. gracilis venom was found to contain 40.3% metalloproteases (SVMPs), 15.3% serine proteases, 6.6% phospholipases A2, 5.0% L-amino acid oxidase, 4.6% Cys-rich secretory proteins (CRISPs), 3.2% disintegrins, 2.9% vascular endothelial growth factors (VEGFs), 1.9% C-type lectin-like proteins, and 20.2% of minor toxins, nontoxins, and unidentified peptides or compounds. Sixteen of these proteoforms matched the toxins whose full amino-acid sequences have been deduced from T. gracilis venom gland cDNA sequences. The hemorrhagic venom of T. gracilis appears to be especially rich in PI-class SVMPs and lacks basic phospholipase A2. We also cloned and sequenced the cDNAs encoding two CRISP and three VEGF variants from T. gracilis venom glands. Sequence alignments and comparison revealed that the PI-SVMP, kallikrein-like proteases, CRISPs, and VEGF-F of T. gracilis and Ovophis okinavensis are structurally most similar, consistent with their close phylogenetic relationship. However, the expression levels of some of their toxins were rather different, possibly due to their distinct ecological and prey conditions.

Keywords: CRISP sequences; Ovophis okinavensis; Trimeresurus gracilis; VEGF sequences; human health; sequence alignments; snake venom proteomics.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Crotalid Venoms* / chemistry
  • Peptides / genetics
  • Phospholipases A2 / chemistry
  • Phospholipases A2 / genetics
  • Phylogeny
  • Proteome / analysis
  • Proteomics / methods
  • Taiwan
  • Trimeresurus*
  • Vascular Endothelial Growth Factor A / genetics

Substances

  • Proteome
  • Vascular Endothelial Growth Factor A
  • Crotalid Venoms
  • Phospholipases A2
  • Peptides

Grants and funding

This research was funded by the Academia Sinica (to I.H.T.), the Ministry of Science and Technology (grant numbers 109-2621-B-020-001, 110-2621-B-020-001), the Council of Agriculture, the Kaohsiung City Government, the National Pingtung University of Science and Technology (grant number NPUST-111005), and the Kaohsiung Chang Gung Memorial Hospital and National Pingtung University of Science and Technology Joint Research Program (grant number CGMH-NPUST-2022-CORPG8M0111).