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| Protein kinase C-beta activates tyrosinase by phosphorylating serine residues in its cytoplasmic domain |
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| Author(s): Park HY, Perez JM, Laursen R, Hara M, Gilchrest BA |
| Source: JOURNAL OF BIOLOGICAL CHEMISTRY Volume: 274 Issue: 23 Pages: 16470-16478 Published: JUN 4 1999 |
| Times Cited: 66 References: 54 |
| Abstract: We have previously shown that protein kinase C-beta (PKC-beta) is required for activation of tyrosinase (Park, H. Y,, Russakovsky, V., Ohno, S., and Gilchrest, B. A. (1993) J. Biol. Chem. 268, 11742-11749), the rate-limiting enzyme in melanogenesis, We now examine its mechanism of activation in human melanocytes. lit vivo phosphorylation experiments revealed that tyrosinase is phosphorylated through the PKC-dependent pathway and that introduction of PKC-beta into nonpigmented human melanoma cells lacking PKC-beta lead to the phosphorylation and activation of tyrosinase. Preincubation of intact melanosomes with purified active PKC-beta in vitro increased tyrosinase activity S-fold. By immunoelectron microscopy, PKC-beta but not PRC-alpha was closely associated with tyrosinase on the outer surface of melanosomes. Western blot analysis confirmed the association of PKC-beta with melanosomes. Only the cytoplasmic (extra-melanosomal) domain of tyrosinase, which contains two serines but no threonines, was phosphorylated by the serine/threonine kinase PKC-beta. These two serines at positions 505 and 509 both are present in the C-terminal peptide generated by trypsin digestion of tyrosinase. Go-migration experiments comparing synthetic peptide standards of all three possible phosphorylated tryptic peptides, a diphosphopeptide and two monophosphopeptides, to tyrosinase-phosphorylated in intact melanocytes by PKC-beta and then subjected to trypsin digestion revealed that both serine residues are phosphorylated by PKC-beta. We conclude that PKC-beta activates tyrosinase directly by phosphorylating serine residues at positions 505 and 509 in the cytoplasmic domain of this melanosome-associated protein. |
| Document Type: Article |
| Language: English |
| Reprint Address: Park, HY (reprint author), Boston Univ, Sch Med, Dept Dermatol, J206,80 E Concord St, Boston, MA 02118 USA |
Addresses:
1. Boston Univ, Sch Med, Dept Dermatol, Boston, MA 02118 USA
2. Boston Univ, Dept Chem, Boston, MA 02118 USA |
| Publisher: AMER SOC BIOCHEMISTRY MOLECULAR BIOLOGY INC, 9650 ROCKVILLE PIKE, BETHESDA, MD 20814 USA |
| Subject Category: Biochemistry & Molecular Biology |
| IDS Number: 202TQ |
| ISSN: 0021-9258 |
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