Comparison of TRPs from murine and human malignant melanocytes.
Most of our knowledge of the mammalian tyrosinase related protein (TRP) activities is derived from studies using murine melanoma models, such as B16 or Cloudman S-91 melanocytes. Owing to the high degree of homology between the murine and human enzymes, it has been assumed that their kinetic behaviour could be similar. However, the protein sequences at the metal binding sites of the murine and human enzymes show some differences of possible functional relevance. These differences are more significant in the metal-A site than in the metal-B site. By using three human melanoma cell lines (HBL, SCL, and BEU), we have studied the catalytic abilities of the human melanogenic enzymes in comparison to those obtained for the counterpart murine enzymes isolated from B16 melanoma. We have found that TRP2 extracted from all cell lines show dopachrome tautomerase activity, although the activity levels in human malignant melanocytes are much lower than in mouse cells. Reconstitution experiments of the human enzyme indicate that TRP2 has Zn at its metal binding-sites. Although mouse tyrosinase does not show DHICA oxidase activity, and this step of the melanogenesis pathway is specifically catalyzed by mouse TRP1, the human enzyme seems to recognize carboxylated indoles. Thus, human tyrosinase could display some residual DHICA oxidase activity, and the function of human TRP1 could differ from that of the murine protein. Attempts to clarify the nature of the metal cofactor in TRP1 were unsuccessful. The enzyme contains mostly Fe and Cu, but the reconstitution of the enzymatic activity from the apoprotein with these ions was not possible.
Martinez-Esparza
M
M
Department of Biochemistry and Molecular Biology, School of Medicine, University of Murcia, Spain.
Jiménez-Cervantes
C
C
García-Borrón
J C
JC
Lozano
J A
JA
del Marmol
V
V
Ghanem
G
G
Solano
F
F
D000818:Animals; D003486:Cyanides / Q000494:pharmacology; D004791:Enzyme Inhibitors / Q000494:pharmacology; D006801:Humans; D007211:Indoles / Q000378:metabolism; D019746:Intramolecular Oxidoreductases; D007535:Isomerases / Q000378:metabolism; D008544:Melanocytes / Q000201:enzymology / Q000378:metabolism*; D008546:Melanoma, Experimental / Q000201:enzymology / Q000378:metabolism*; D008562:Membrane Glycoproteins; D051379:Mice; D014442:Monophenol Monooxygenase / Q000037:antagonists & inhibitors / Q000378:metabolism; D010088:Oxidoreductases; D011506:Proteins / Q000378:metabolism*; D014407:Tumor Cells, Cultured
D003486:Cyanides; D004791:Enzyme Inhibitors; D007211:Indoles; D008562:Membrane Glycoproteins; D011506:Proteins; D010088:Oxidoreductases; C489799:TYRP1 protein, human; C079621:Tyrp1 protein, mouse; C064056:tyrosinase-related protein-1; D014442:Monophenol Monooxygenase; D007535:Isomerases; D019746:Intramolecular Oxidoreductases; C042501:dopachrome isomerase