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© Research
Publication : European journal of biochemistry / FEBS

Identification and characterization of a melanocyte-specific novel 65-kDa peripheral membrane protein

Scientific Fields
Diseases
Organisms
Applications
Technique

Published in European journal of biochemistry / FEBS - 01 Dec 1999

Samaraweera P, Donatien PD, Qazi S, Kobayashi T, Hearing VJ, Panthier JJ, Orlow SJ

Link to Pubmed [PMID] – 10583387

Eur. J. Biochem. 1999 Dec;266(3):924-34

In order to study proteins of the melanosome, we developed a panel of antisera against various protein fractions of melanosomes from B16 melanoma cells. An antiserum raised against a Triton X-100 insoluble fraction of melanosomes recognized a 65-kDa protein in melanocytes from mice homozygous for the buff mutation, but not in their wild type counterparts. Further studies were conducted using a specific, second generation antiserum raised against the purified protein. The protein was also detected in melanocytes cultured from albino mice, but absent in cultured mouse cell lines not of melanocyte origin. Density gradient centrifugation of subcellular organelles and indirect immunofluorescent cell staining, indicated that the protein was associated with melanosomes and vesicles. The protein on intact organelles could be made soluble using sodium carbonate, and digested with proteases in the absence of detergent suggesting that it was a peripheral membrane protein localized on the cytosolic face of organelle membranes. Metabolic labelling of cells and N-glycosidase F digestion of cell extracts indicated that the protein was not N-glycosylated. Based on its intracellular localization and biochemical defects in the buff mouse, a potential role has been suggested for the 65-kDa protein in intracellular membrane trafficking.

http://www.ncbi.nlm.nih.gov/pubmed/10583387