Specificity of Loxosceles α clade phospholipase D enzymes for choline-containing lipids: Role of a conserved aromatic cage

Emmanuel E. Moutoussamy, Qaiser Waheed, Greta J. Binford, Hanif M. Khan, Shane M. Moran, Anna R. Eitel, Matthew H.J. Cordes, Nathalie Reuter

Research output: Contribution to journalArticlepeer-review


Spider venom GDPD-like phospholipases D (SicTox) have been identified to be one of the major toxins in recluse spider venom. They are divided into two major clades: The α clade and the β clade. Most α clade toxins present high activity against lipids with choline head groups such as sphingomyelin, while activities in β clade toxins vary and include preference for substrates containing ethanolamine headgroups (Sicarius terrosus, St_βIB1). A structural comparison of available structures of phospholipases D (PLDs) reveals a conserved aromatic cage in the α clade. To test the potential influence of the aromatic cage on membrane- lipid specificity we performed molecular-dynamics (MD) simulations of the binding of several PLDs onto lipid bilayers containing choline headgroups; two SicTox from the α clade, Loxosceles intermedia αIA1 (Li_αIA) and Loxosceles laeta αIII1 (Ll_αIII1), and one from the β clade, St_βIB1. The simulation results reveal that the aromatic cage captures a choline-headgroup and suggest that the cage plays a major role in lipid specificity. We also simulated an engineered St_βIB1, where we introduced the aromatic cage, and this led to binding with choline-containing lipids. Moreover, a multiple sequence alignment revealed the conservation of the aromatic cage among the α clade PLDs. Here, we confirmed that the i-face of α and β clade PLDs is involved in their binding to choline and ethanolamine-containing bilayers, respectively. Furthermore, our results suggest a major role in choline lipid recognition of the aromatic cage of the α clade PLDs. The MD simulation results are supported by in vitro liposome binding assay experiments.

Original languageEnglish (US)
Article numbere1009871
JournalPLoS computational biology
Issue number2
StatePublished - Feb 2022

ASJC Scopus subject areas

  • Ecology, Evolution, Behavior and Systematics
  • Ecology
  • Modeling and Simulation
  • Molecular Biology
  • Genetics
  • Cellular and Molecular Neuroscience
  • Computational Theory and Mathematics


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