メラニン生成機構にかんする2，3の問題

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The recent availability of cell biochemical techniques as ultracentrifugation separation of cell particles and density gradient centrifugation made possible the microdissection of and biochemical studies on melanin-forming cells which have elucidated the steps involved in the formation of melanosomes. A series of experiments along the studies on melanogenesis has been carried out within the last several years. Experimental data obtained from these studies are all compartible with the hypothesis, melanosome concept that tyrosinase is synthesized in ribosomes, transferred via endoplasmic reticulum to the Golgi area, where tyrosinase is separated into small units, each of which is surrounded by a membranous envelope. Within each envelope, the tyrosinase molecules assume an ordered pattern, after which melanin biosynthesis begins and the particle is known as a melanosome. As melanin gradually accumulates, the melanosome is transformed into a uniformly dense and structureless particle, a mature melanosome. The problems described herein are the in vitro experiments on the melanization process of melanosomes and the controling mechanism of thiol-group in melanin formation in melanocyte. The reciprocal relationship between tyrosinase activity and melanization of melanosomes has been demonstrated and data presented herein would be additional evidence to support this concept. Two kinds of melanosomes in defferent developmental stages were prepared from the mouse melanoma. Melanosome No. 1 is less melanized and possesses more tyrosinase activity than melanosome No. 2. Comparison of the rate of incorporation of 14C-DOPA in vitro by melanosome No. 1 and No. 2 was carried out. The experimental results showed that melanosome No. 1, immature melanosome can synthesize and deposit more melanin than melanosome No. 2, mature melanosomes. The melanization mechanism of melanosomes was further studied. Melanosomes isolated from the mouse melanoma were incubated with the excess amount of 14C-DOPA. The amount of melanin formed was dependent on the amount of melanosomes incubated but relationship between them was an upward convex rather than a straight line. It was assumed that there are some kinds of mechanisms which inactivate the enzyme. Kinetic studies were carried out in order to analyze the kind of inactivation mechanisms thought to be present. The exprimental results obtained could not be explained by assuming only one type of inactive enzymeproduct complex. However, they could be explained on the basis of simultaneous formation of two or more types of inactive enzymn-product complex. The sulfhydryl group has been considered as one of the important controling factors in melanin formation in melanocytes. The inhibitory effects of thiol compounds ont yrosine hydroxylase and tyrosinase in melanosomes isolated from mouse melanoma are descrided and an in vitro study of the reaction between an oxidation product of DOPA and glutathione in the presence of mammalian tyrosinase is reported. The formation of new chemical compounds resulted from the presence of DOPA, glutathione and mammalian tyrosinase in solution at pH 6.8. The existence of chemical bonding between glutathione and an oxidation product of DOPA, probably DOPA-quinone was demonstrated. The glutathione-derived reaction product remained soluble in the reaction medium and did not precipitate on the melanosomes. The pKi values of glutathione and cysteine on tyrosine hydroxylase were much smaller than those on tyrosinase. The inhibition of tyrosine melanin formation by sulfhydryl compounds may take place in two ways: by inhibition of the enzyme tyrosinase, and by the formation of new compounds in which intermediates of the tyrosine-tyrosinase reaction are trapped by the sulfhydryl compounds. (to be cotinued)