A new mechanism for metal ion-assisted interchain helix assembly in a naturally occurring peptide mediated by optimally spaced gamma-carboxyglutamic acid residues.
Helix-helix interactions, such as those that occur in coiled-coil domains, four-helix bundles, or membrane-spanning helical bundles, are important to the structural organization and function of numerous proteins. However, tractable peptide models for studying such structural elements have been limited to synthetic analogs of coiled-coil protein domains and de novo designed peptides. The present study provides evidence that conantokin-G (con-G), a gamma-carboxyglutamate (Gla)-rich neuroactive peptide from a venomous marine snail, can self-associate in the presence of certain divalent metal cations. Sedimentation equilibrium analyses of con-G show that Ca2+ binding promotes peptide dimerization, while the addition of the tighter binding divalent cations, Mg2+, Zn2+, and Mn2+, does not result in intermolecular association. The effects of specific residue replacements indicate that an i, i + 4, i + 7, i + 11 arrangement of Gla residues is essential for con-G self-assembly. To determine the relative chain orientation of the dimeric assembly, distributions of Cys-containing con-G variants were examined in thiol-disulfide rearrangement assays and the results were consistent with an antiparallel alignment. Our data suggest that the driving force for con-G dimerization stems from the appropriate balance of interchain and intrachain metal ion coordination by Gla residues in similar locations. These findings suggest a new role for Gla residues and accompanying cation binding in the stabilization of interstrand helix association in a natural product and provide a model for controlled assembly of peptide chains or segments of larger proteins.
Dai
Qiuyun
Q
Department of Chemistry and Biochemistry and the W. M. Keck Center for Transgene Research, University of Notre Dame, Notre Dame, IN 46556, USA.
Prorok
Mary
M
Castellino
Francis J
FJ
D015055:1-Carboxyglutamic Acid / Q000737:chemistry*; D000595:Amino Acid Sequence; D000818:Animals; D002151:Calorimetry; D002413:Cations, Divalent / Q000737:chemistry; D002942:Circular Dichroism; D020916:Conotoxins / Q000737:chemistry* / Q000235:genetics; D004220:Disulfides; D018691:Excitatory Amino Acid Antagonists / Q000737:chemistry; D008670:Metals / Q000737:chemistry*; D008978:Mollusk Venoms / Q000737:chemistry; D010455:Peptides / Q000737:chemistry* / Q000235:genetics; D017433:Protein Structure, Secondary
D002413:Cations, Divalent; D020916:Conotoxins; D004220:Disulfides; D018691:Excitatory Amino Acid Antagonists; D008670:Metals; D008978:Mollusk Venoms; D010455:Peptides; D015055:1-Carboxyglutamic Acid; C055407:conotoxin GV