Aspartic Acid Isomerization Characterized by High Definition Mass Spectrometry Significantly Alters the Bioactivity of a Novel Toxin from Poecilotheria

Toxins (Basel). 2020 Mar 25;12(4):207. doi: 10.3390/toxins12040207.

Abstract

Research in toxinology has created a pharmacological paradox. With an estimated 220,000 venomous animals worldwide, the study of peptidyl toxins provides a vast number of effector molecules. However, due to the complexity of the protein-protein interactions, there are fewer than ten venom-derived molecules on the market. Structural characterization and identification of post-translational modifications are essential to develop biological lead structures into pharmaceuticals. Utilizing advancements in mass spectrometry, we have created a high definition approach that fuses conventional high-resolution MS-MS with ion mobility spectrometry (HDMSE) to elucidate these primary structure characteristics. We investigated venom from ten species of "tiger" spider (Genus: Poecilotheria) and discovered they contain isobaric conformers originating from non-enzymatic Asp isomerization. One conformer pair conserved in five of ten species examined, denominated PcaTX-1a and PcaTX-1b, was found to be a 36-residue peptide with a cysteine knot, an amidated C-terminus, and isoAsp33Asp substitution. Although the isomerization of Asp has been implicated in many pathologies, this is the first characterization of Asp isomerization in a toxin and demonstrates the isomerized product's diminished physiological effects. This study establishes the value of a HDMSE approach to toxin screening and characterization.

Keywords: Poecilotheria; Voltage-gated sodium channel; deamidation; high definition mass spectrometry; ion mobility; isomerization; poecilotheriatoxin; supplemental activation; venom.

MeSH terms

  • Amino Acid Sequence
  • Animals
  • Aspartic Acid / chemistry*
  • Binding Sites
  • Cell Line
  • Drug Discovery
  • Humans
  • Ion Mobility Spectrometry*
  • Isomerism
  • Mass Spectrometry*
  • Membrane Potentials
  • NAV1.7 Voltage-Gated Sodium Channel / drug effects*
  • NAV1.7 Voltage-Gated Sodium Channel / metabolism
  • Neurotoxins / chemistry
  • Neurotoxins / pharmacology*
  • Protein Binding
  • Protein Conformation
  • Protein Processing, Post-Translational
  • Spider Venoms / chemistry
  • Spider Venoms / pharmacology*
  • Structure-Activity Relationship
  • Voltage-Gated Sodium Channel Agonists / chemistry
  • Voltage-Gated Sodium Channel Agonists / pharmacology*

Substances

  • NAV1.7 Voltage-Gated Sodium Channel
  • Neurotoxins
  • Spider Venoms
  • Voltage-Gated Sodium Channel Agonists
  • Aspartic Acid