The receptor for alpha-melanotropin of mouse and human melanoma cells. Application of a potent alpha-melanotropin photoaffinity label.

Abstract

The melanotropin (MSH) receptor of mouse B16-F1 melanoma cells was characterized by photoaffinity cross-linking, using a potent alpha-MSH photolabel, [norleucine4, D-phenylalanine7, 1'-(2-nitro-4-azidophenylsulfenyl)-tryptophan9]-alpha-melanotropin (Naps-MSH). Its monoiodinated form, 125I-Naps-MSH, displayed a approximately 6.5-fold higher biological activity than alpha-MSH. Scatchard analysis of the saturation curves with 125I-Naps-MSH revealed approximately 20,000 receptors/B16-F1 cell and an apparent KD of approximately 0.3 nM. Analysis of the cross-linked MSH receptor by sodium dodecyl sulfate-polyacrylamide gel electrophoresis showed that a photolabeled band of approximately 45 kDa occurs in B16-F1, B16-F10, and Cloudman S91 mouse melanoma, as well as in human D10 and 205 melanoma but not in non-melanoma cells. The labeled 45-kDa protein had an isoelectric point of 4.5-4.9 as determined by two-dimensional gel electrophoresis. Treatment of the labeled 45-kDa protein of B16-F1 cell membranes by neuraminidase shifted the band to approximately 42 kDa. A similar band of about 42 kDa was also observed after receptor labeling of B16-W4 cells, a cell line with a decreased number of terminal N-linked neuraminyl residues. These results indicate that the labeled 45-kDa glycoprotein contains terminal sialic acid residues, explaining the low pI of this protein, and that it is characteristic for melanoma cells and hence part of the MSH receptor.

• Authors: F Solca, W Siegrist, R Drozdz, J Girard, A N Eberle
• Journal: The Journal of biological chemistry (J Biol Chem) Vol. 264 Issue 24 Pg. 14277-81 (Aug 25 1989) ISSN: 0021-9258 [Print] United States
• PMID: 2547792 (Publication Type: Journal Article, Research Support, Non-U.S. Gov't)

Chemical References

• Affinity Labels
• Glycoproteins
• Receptors, Pituitary Hormone
• alpha-MSH
• MSH receptor
• Neuraminidase

Topics

• Affinity Labels
• Animals
• Cell Line
• Glycoproteins (isolation & purification, metabolism)
• Humans
• Isoelectric Point
• Melanoma (analysis, metabolism)
• Mice
• Molecular Weight
• Neuraminidase
• Photochemistry
• Rats
• Receptors, Pituitary Hormone (analysis, isolation & purification)
• Tumor Cells, Cultured (analysis, metabolism)
• alpha-MSH (metabolism)