Subcellular distribution of glycosidases in human polymorphonuclear leucocytes.
The subcellular distribution of nine glycosidases were studied in fractions of homogenized human polymorphonuclear leucocytes (neutrophils) obtained by isopycnic centrifugation through linear sucrose density gradients. The substrates were 4-methylumbelliferyl glycosides. All nine glycosides were hydrolysed by enzymes in neutrophil cytosol fractions, and by enzymes in at least one granule population. alpha-Glucosidase activity sedimented in sucrose density gradients to a point (p = 1.180 g/ml) just above the specific granules, possibly the 'tertiary' granule population. The peak corresponding to alpha-glucosidase did not co-sediment with, but considerably overlapped, the peak corresponding to lactoferrin, a marker for specific granules (p = 1.187 g/ml). alpha-Galactosidase activity was found primarily in heavy azurophil granules (p = 1.222 g/ml). alpha-Mannosidase activity was found primarily in light azurophil granules (p = 1.206 g/ml), following the distribution of myeloperoxidase, the commonly used azurophil granule marker. beta-Glucosidase activity was concentrated in mitochondrial fractions (p = 1.160 g/ml). All other glycosidases presented complex distributions, with activities not restricted to one granule class. Granule-associated glycosidase activities were increased 2--38 times when measured in the presence of 0.05% Triton X-100, indicating latency of the enzymes within granules.
Subcellular distribution of glycosidases in human polymorphonuclear leucocytes. The subcellular distribution of nine glycosidases were studied in fractions of homogenized human polymorphonuclear leucocytes (neutrophils) obtained by isopycnic centrifugation through linear sucrose density gradients. The substrates were 4-methylumbelliferyl glycosides. All nine glycosides were hydrolysed by enzymes in neutrophil cytosol fractions, and by enzymes in at least one granule population. alpha-Glucosidase activity sedimented in sucrose density gradients to a point (p = 1.180 g/ml) just above the specific granules, possibly the 'tertiary' granule population. The peak corresponding to alpha-glucosidase did not co-sediment with, but considerably overlapped, the peak corresponding to lactoferrin, a marker for specific granules (p = 1.187 g/ml). alpha-Galactosidase activity was found primarily in heavy azurophil granules (p = 1.222 g/ml). alpha-Mannosidase activity was found primarily in light azurophil granules (p = 1.206 g/ml), following the distribution of myeloperoxidase, the commonly used azurophil granule marker. beta-Glucosidase activity was concentrated in mitochondrial fractions (p = 1.160 g/ml). All other glycosidases presented complex distributions, with activities not restricted to one granule class. Granule-associated glycosidase activities were increased 2--38 times when measured in the presence of 0.05% Triton X-100, indicating latency of the enzymes within granules.