Studies on cutin-esterase II. Characteristics of cutin-esterase from Botrytis cinerea and its activity on tomato-cutin

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The activity of cutin-esterase, cutinase, was detected in the mycelial homogenate of Bolrytis cinerea cultured in a peptone-sucrose medium at 25℃ for 7 days. The crude enzyme solution was prepared from the homogenate by centrifugation at 106,600×g, treatment with (NH_4)_2SO_4 at 70% saturation, and dialysis against 0.01_M phosphate buffer. The optimum pH, temperature and assay duration for enzyme activity were 5.0, 25℃ and 18 hr, respectively. Specific activity was 255 mμmoles/mg protein/18 hr as palmitate under optimum conditions. 830% of the activity was lost by heating the enzyme solution (pH 7.6) for 4 min at 95℃. Palmitic, stearic, oleic, 9,10-dihydroxystearic or linoleic, dihydroxyeicosanoic and octadecanedioic acids were recognized in the enzymic hydrolysate of tomato-cutin using gas-liquid chromatography. Among these fatty acids, palmitic, oleic and octadecanedioic acids were readily liberated by the enzyme, but dihydroxyeicosanoic acid, the major component of tomato-cutin, was isolated only in small amounts. The enzyme is, therefore, an exo-type cutinase which hydrolyses minor side chains of fatty acids bound to the major structure of cutin. Cutinesterase may facilitate cuticular invasion by fungi as a result of reduction in mechanical strength of the cuticle by the enzyme.
日本植物生理学会の論文