The three-dimensional structure of an H-superfamily conotoxin reveals a granulin fold arising from a common ICK cysteine framework

J Biol Chem. 2019 May 31;294(22):8745-8759. doi: 10.1074/jbc.RA119.007491. Epub 2019 Apr 11.

Abstract

Venomous marine cone snails produce peptide toxins (conotoxins) that bind ion channels and receptors with high specificity and therefore are important pharmacological tools. Conotoxins contain conserved cysteine residues that form disulfide bonds that stabilize their structures. To gain structural insight into the large, yet poorly characterized conotoxin H-superfamily, we used NMR and CD spectroscopy along with MS-based analyses to investigate H-Vc7.2 from Conus victoriae, a peptide with a VI/VII cysteine framework. This framework has CysI-CysIV/CysII-CysV/CysIII-CysVI connectivities, which have invariably been associated with the inhibitor cystine knot (ICK) fold. However, the solution structure of recombinantly expressed and purified H-Vc7.2 revealed that although it displays the expected cysteine connectivities, H-Vc7.2 adopts a different fold consisting of two stacked β-hairpins with opposing β-strands connected by two parallel disulfide bonds, a structure homologous to the N-terminal region of the human granulin protein. Using structural comparisons, we subsequently identified several toxins and nontoxin proteins with this "mini-granulin" fold. These findings raise fundamental questions concerning sequence-structure relationships within peptides and proteins and the key determinants that specify a given fold.

Keywords: CD spectroscopy; NMR spectroscopy; antistasin; conotoxin; disulfide; disulfide bond; granulin; inhibitor cystine knot; protein conformation; protein evolution; protein expression; protein structure; protein-disulfide isomerase; toxin; β-hairpin.

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, Non-U.S. Gov't

MeSH terms

  • Amino Acid Sequence
  • Animals
  • Conotoxins / chemistry*
  • Conotoxins / genetics
  • Conotoxins / metabolism
  • Conus Snail / metabolism*
  • Cysteine / chemistry*
  • Disulfides / chemistry
  • Granulins / chemistry*
  • Granulins / metabolism
  • Magnetic Resonance Spectroscopy
  • Mollusk Venoms / metabolism
  • Protein Conformation, beta-Strand
  • Protein Folding
  • Protein Stability
  • Recombinant Proteins / biosynthesis
  • Recombinant Proteins / chemistry
  • Recombinant Proteins / genetics

Substances

  • Conotoxins
  • Disulfides
  • Granulins
  • Mollusk Venoms
  • Recombinant Proteins
  • Cysteine

Associated data

  • PDB/1QK7
  • PDB/2JYE
  • PDB/4X9Z
  • PDB/1C9T
  • PDB/2KCX
  • PDB/2K2C
  • PDB/6Q5Z