Hiroo Sanada

Effect of thiol compounds on melanin formation by tyrosinase

Abstract 1. 1. The inhibitory effect of sulfhydryl compounds on the tyrosinase-dependent melanin formation was studied. 2. 2. In the presence of cysteine, pyrocatechol was conjugated with it by tyrosinase (EC. 1.10.3.1) action to produce a colorless material. This adduct was found to be S -(2,3-dihydroxyphenyl)cysteine. 3. 3. The experiments with L -[2,5,6- 3 H 3 ]DOPA and l -[3,5- 3 H 2 ]tyrosine indicated that the hydrogen atom at C-5 of DOPA was substituted by thiol groups, this reaction was in agreement with the formation of the pyrocatechol-cysteine conjugate. 4. 4. The pyrocatechol-cysteine conjugate was also obtained by the non-enzymatic conjugation of o -quinone with cysteine. This fact indicated that tyrosinase only oxidized catechol to o -quinone which then combined with sulfhydryl groups non-enzymatically. 5. 5. These conjugation products were not oxidized further by tyrosinase, but gave some inhibitory effects on the melanin formation by the enzyme.

A defect of purine nucleotide cycle in the skeletal muscle of hereditary dystrophic mice.

Abstract The purine nucleotide cycle in the hind leg skeletal muscle of hereditary dystrophic mice ( C57BL 6J - dy dy ) was investigated. The amount of adenine nucleotide produced from adenylosuccinate by the muscle extract in the dystrophic group was less than 3 % of that in the control group, while adenine nucleotide plus adenylosuccinate converted from IMP in the dystrophic group was about 70 % of that of the control group. Moreover, the activity of AMP deaminase of the dystrophic group was about 50 % of that of the control group. These results indicate that the purine nucleotide cycle is defective in the dystrophic muscle. This abnormality was suggested to be caused by the considerably low activity of adenylosuccinase.

Effect of ascorbic acid on tyrosine hydroxylase activity in vivo

Abstract The activity of tyrosine hydroxylase in the homogenate of adrenal gland decreased in scurvy, and it was recovered by the administration of ascorbic acid. The mechanism of increase in tyrosine hydroxylase activity by administration of ascorbic acid has been studied. The enzyme activities of the adrenal homogenates in nonscorbutic and scorbutic guinea pigs were changed neither by dialysis nor by gel filtration on Sephadex G-25. However, stimulation of enzyme activity by the administration of ascorbic acid, was blocked either by puromycin or by actinomycin D. Tyrosine hydroxylase was purified by ammonium sulfate fractionation, Sephadex G-200, and hydroxylapatite chromatography. Antibody to the partially purified enzyme was prepared in rabbit. Immunochemical analysis indicated that there was a constant amount of immunochemically precipitable enzyme per unit of enzyme activity. The studies reported here showed that the increase of enzyme activity by the administration of ascorbic acid was due to the increased amount of the enzyme protein.

Induction of tyrosine hydroxylase by dibutyryl adenosine 3′,5′-monophosphate in guinea pig adrenal slices

Abstract The activity of tyrosine hydroxylase was induced by dibutyryl adenosine 3′, 5′-monophosphate (dibutyryl cyclic AMP) in slices of guinea pig adrenal glands. One hour after the addition of dibutyryl cyclic AMP, the enzyme level started to increase and reached the maximum by 2 hr. Both cycloheximide and actinomycin D largely prevented the effect of dibutyryl cyclic AMP on the enzyme. When the slice was incubated without bovine serum, the enzyme activity was not stimulated by dibutyryl cyclic AMP. Therefore, the effect of dibutyryl cyclic AMP on tyrosine hydroxylase was found to be due to an increased rate of enzyme synthesis. These findings support the view that dibutyryl cyclic AMP is an inducer of tyrosine hydroxylase in adrenal glands of guinea pig.