Molecular structure and granulocyte/macrophage colony-stimulating factor activity.
Granulocyte/macrophage colony-stimulating factor (GM-CSF) plays a critical role in myeloid differentiation and in several immune and inflammatory processes. GM-CSF binds to specific cellular receptors (GM-CSFR) which belong to a recently described supergene family. These receptors are potential targets for pharmacologic design and such design depends on a molecular understanding of ligand-receptor interactions. We present our initial studies evaluating the potential active sites of the molecule. The sites on the GM-CSF molecule that were studied represent two alpha-helices predicted to be critical for GM-CSF activity, as implicated by human-murine chimeric molecule studies. These helices are predicted to be adjacent in native GM-CSF. Peptides corresponding to amino acids 17-31 and 78-99 of GM-CSF were synthesized and cross-linked to one another in two different orientations. The ability of anti-GM-CSF to bind the individual and complexed peptides was evaluated by both ELISA and radioimmunoassay. Significant binding to all peptides was demonstrated. A preferred orientation of the two peptides was apparent, and this agreed with the predicted model structures. Antibodies were developed against the coupled peptides, and these demonstrated significant cross-reactivity with recombinant human GM-CSF. Additionally, analyses of anti-peptide antisera binding studies predict these two amino acid sequences to lie in parallel planes to one another in the native human GM-CSF molecule.
Von Feldt
J M
JM
Department of Medicine, University of Pennsylvania School of Medicine, Philadelphia 19104.
Kieber-Emmons
T
T
Weiner
D B
DB
Ugen
K E
KE
Williams
W V
WV
D000595:Amino Acid Sequence; D001665:Binding Sites; D004797:Enzyme-Linked Immunosorbent Assay; D000939:Epitopes / Q000737:chemistry / Q000276:immunology / Q000378:metabolism; D016178:Granulocyte-Macrophage Colony-Stimulating Factor / Q000737:chemistry / Q000276:immunology / Q000378:metabolism*; D006801:Humans; D007106:Immune Sera; D008969:Molecular Sequence Data; D010446:Peptide Fragments / Q000235:genetics / Q000276:immunology / Q000378:metabolism; D011487:Protein Conformation; D011863:Radioimmunoassay
D000939:Epitopes; D007106:Immune Sera; D010446:Peptide Fragments; D016178:Granulocyte-Macrophage Colony-Stimulating Factor