A new approach to the measurement of phospholipase A2 in tissue homogenates and its application to human skin.
We describe a new approach to the quantification of tissue phospholipase A2 based on the observation that the Vmax is increased by several orders of magnitude in the presence of high concentrations of dimethyl sulphoxide. The enzyme from human skin showed optimal activity at 30% dimethyl sulphoxide and pH 6-7.5, and had an absolute dependence on Ca2+ (apparent Km = 1.4 mmol/l). The relationship between reaction velocity and substrate concentration did not follow Michaelis-Menten kinetics. The enzyme was inhibited by p-bromophenacyl bromide, half-maximal inhibition occurring at 2.4 mumol/l. Surprisingly, assay of specimens of various human tissues indicated that, with the exception of pancreas, skin contained higher levels of phospholipase A2 activity than any other tissue investigated. Equally surprising was that primate skin (in particular human) seems considerably more active than that of other mammals. Speculatively, these findings may be related to the high levels of free fatty acids occurring on the skin surface.
D000818:Animals; D002118:Calcium / Q000494:pharmacology; D002250:Carbon Radioisotopes; D004121:Dimethyl Sulfoxide / Q000494:pharmacology*; D006801:Humans; D007700:Kinetics; D009928:Organ Specificity; D010740:Phospholipases / Q000378:metabolism*; D010741:Phospholipases A / Q000378:metabolism*; D054467:Phospholipases A2; D011865:Radioisotope Dilution Technique; D012867:Skin / Q000201:enzymology*; D013045:Species Specificity
D002250:Carbon Radioisotopes; D010740:Phospholipases; D010741:Phospholipases A; D054467:Phospholipases A2; D002118:Calcium; D004121:Dimethyl Sulfoxide