Topology of the calmodulin-melittin complex

Andrea Scaloni, Nadia Miraglia, Stefania Orrù, Pietro Amodeo, Andrea Motta, Gennaro Marino, Piero Pucci

Research output: Contribution to journalArticle

Abstract

The topology of the Ca2+-calmodulin-melittin ternary complex has been investigated by a combined strategy which integrates limited proteolysis and cross-linking experiments with mass spectrometric methodologies. The rationale behind the methods is that the interface regions of two interacting proteins are accessible to the solvent in the isolated molecules, whereas they become protected following the formation of the complex. Therefore, when limited proteolysis experiments are carried out on both the isolated proteins and the complex, differential peptide maps are obtained from which the interface regions can be inferred. Alternatively, cross-linking reactions performed under strictly controlled conditions lead to the identification of spatially closed amino acid residues in the complex. Mass spectrometry can be employed in both procedures for the definition of the cleavage sites and to identify covalently linked residues. Our results show that melittin interacts with calmodulin by adopting a parallel orientation, i.e. the N and C-terminal halves of the peptide are anchored to the amino and carboxy-terminal domains of the protein, respectively. This orientation is inverted with respect to all the peptide substrates examined so far. A model of the complex was designed and refined on the basis of the experimental results, supporting the above conclusions. This finding reveals a further dimension to the already remarkable capability of calmodulin in binding different protein substrates, providing this protein with the capability of regulating an even larger number of enzymes.

Original languageEnglish
Pages (from-to)945-958
Number of pages14
JournalJournal of Molecular Biology
Volume277
Issue number4
DOIs
Publication statusPublished - Apr 10 1998

Keywords

  • Calmodulin
  • Cross-links
  • Limited proteolysis
  • Mass spectrometry
  • Protein-protein interaction

ASJC Scopus subject areas

  • Virology

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  • Cite this

    Scaloni, A., Miraglia, N., Orrù, S., Amodeo, P., Motta, A., Marino, G., & Pucci, P. (1998). Topology of the calmodulin-melittin complex. Journal of Molecular Biology, 277(4), 945-958. https://doi.org/10.1006/jmbi.1998.1629