1.Acid lipase activity of human lymphocytes.
Coates PM, Cortner JA, Hoffman GM, Brown SA. Biochim Biophys Acta. 1979 Feb 26;572(2):225-34.
Acid lipase activity was examined in human leukocytes using 4-methylumbelliferyl esters in a fluorimetric assay. Mononuclear leukocytes had 10--15 times the acid lipase activity of polymorphonuclear leukocytes. The enzyme activity was highest using the oleate ester of 4-methylumbelliferone at pH 4.0, in the presence of L-alpha-phosphatidylcholine and taurodeoxycholic acid (sodium salt). Acid lipase activity was inhibited by diethylaminoethoxyhexestrol, sodium chloride and fluoride, potassium chloride, calcium chloride and albumin, but was unaffected by diethyl p-nitrophenyl phosphate or sulphydryl reagents. There were at least two forms of acid lipase activity: one (A form) was sensitive to heart inactivation (56 degrees C) and corresponded to the enzyme deficient in patients with Wolman's disease; the other (B form) was resistant to heat inactivation and corresponded to the residual enzyme activity found in Wolman's disease.
2.A photoreactive competitive inhibitor of the human lysosomal neuraminidase in cultured skin fibroblasts.
Warner TG1. Biochem Biophys Res Commun. 1987 Nov 13;148(3):1323-9.
A photoreactive, potent, competitive inhibitor of the human lysosomal neuraminidase in cultured skin fibroblasts has been prepared. The starting material, 2,3 dehydro-N-acetyl neuraminic acid methyl ester, was selectively tosylated at the C-9 position with tosyl chloride and subsequently peracetylated with acetic anhydride. The tosyl group was displaced with potassium thio acetate in dimethylformamide at 60 degrees C for 80 min. 4-fluoro-3-nitrophenylazide was incorporated by reaction with the thio acetate product and equimolar sodium methoxide in methanol followed by reacetylation. Base hydrolysis gave the final product, 9-S-(4-azido-2-nitrophenyl)-5-acetamido-2,6 anhydro-2,3,5,9-tetradeoxy-9-thio-D-glycero-D-galacto-non-2-enonic acid (W5). The yields at each step were 50-70%. Competitive inhibition kinetics were observed when W5 was tested with the fibroblast neuraminidase using 4-methylumbelliferyl-N-acetyl-neuraminic acid as substrate giving an apparent Ki of about 10 microM.