Neutrophil-activating peptide-2 and melanoma growth-stimulatory activity are functional as monomers for neutrophil activation

Krishna Rajarathnam, Cyril M. Kay, Beatrice Dewald, Marlene Wolf, Marco Baggiolini, Ian Clark-Lewis, Brian D. Sykes

Research output: Contribution to journalArticle

40 Scopus citations

Abstract

Neutrophil-activating peptide-2 (NAP-2) and melanoma growth-stimulatory activity (MGSA) are members of the chemokine family of inflammatory proteins. The structures of NAP-2, determined by x-ray crystallography, and MGSA, elucidated by NMR spectroscopy, revealed a tetramer and dimer, respectively. In order to address the relevance of multimeric species to their activities on neutrophils, analogs of NAP-2 and MGSA were synthesized in which the backbone amide proton of Leu-22 in NAP-2, and Val-26 in MGSA, was substituted with the bulky methyl group (NH → NCH3). These analogs were shown to be monomeric by sedimentation equilibrium ultracentrifugation studies and were similar to the corresponding native protein in assays for neutrophil elastase release and Ca2+ mobilization from IL-8R1 and IL-8R2 transformed cells. Sedimentation equilibrium studies of the native NAP-2 and MGSA were also carried out to address the association behavior. For NAP-2, there was no evidence for the tetramer, but an equilibrium between monomers and dimers and the dissociation constant was calculated to be 50-100 μM. Similarly, MGSA showed a monomer-dimer equilibrium with a K(d) of ~5 μM. The data from the monomeric analogs and also the calculation of dissociation constants indicate that NAP-2 and MGSA have a tendency to associate above the concentrations required for maximal activity or for receptor activation, but at functional concentrations they are predominantly monomers.

Original languageEnglish (US)
Pages (from-to)1725-1729
Number of pages5
JournalJournal of Biological Chemistry
Volume272
Issue number3
DOIs
StatePublished - Jan 1 1997
Externally publishedYes

ASJC Scopus subject areas

  • Biochemistry
  • Molecular Biology
  • Cell Biology

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