Membrane anchoring and surface distribution of glycohydrolases of human erythrocyte membranes.
The membrane anchoring of the following glycohydrolases of human erythrocyte plasma membranes was investigated: alpha- and beta-D-glucosidase, alpha- and beta-D-galactosidase, beta-D-glucuronidase, N-acetyl-beta-D-glucosaminidase, alpha-D-mannosidase, and alpha-L-fucosidase. Optimized fluorimetric methods for the assay of these enzymes were set up. Treatment of the ghost preparation with 1.0 mol/l (optimal concentration) NaCl caused release ranging from 4.2% of alpha-D-glucosidase to 70% of beta-D-galactosidase; treatment with 0.4% (optimal concentration) Triton X-100 liberated 5.1% of beta-D-galactosidase to 89% of alpha-D-glucosidase; treatment with 1.75% (optimal concentration) octylglucoside yielded solubilization from 6.3% of beta-D-galactosidase to 85% of alpha-D-glucosidase. Treatment with phosphoinositide-specific phospholipase C caused no liberation of any of the studied glycohydrolases. These results are consistent with the notion that the above glycohydrolases are differently anchored or associated with the erythrocyte plasma membrane, and provide the methodological basis for inspecting the occurrence of these enzymes in different membrane microdomains.
Goi
G
G
Department of Medical Chemistry and Biochemistry, The Medical School, University of Milan, Via Saldini 50, 20133, Milan, Italy.
Bairati
C
C
Massaccesi
L
L
Lovagnini
A
A
Lombardo
A
A
Tettamanti
G
G
D000328:Adult; D003902:Detergents / Q000494:pharmacology; D004910:Erythrocyte Membrane / Q000187:drug effects / Q000201:enzymology* / Q000378:metabolism*; D005260:Female; D005470:Fluorometry; D005960:Glucosides / Q000494:pharmacology; D006026:Glycoside Hydrolases / Q000032:analysis / Q000378:metabolism*; D017261:Glycosylphosphatidylinositols / Q000378:metabolism; D006801:Humans; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D008297:Male; D008565:Membrane Proteins / Q000032:analysis / Q000378:metabolism*; D008875:Middle Aged; D017830:Octoxynol / Q000494:pharmacology; D009994:Osmolar Concentration; D043265:Phosphatidylinositol Diacylglycerol-Lyase; D012965:Sodium Chloride / Q000494:pharmacology; D012995:Solubility / Q000187:drug effects; D012996:Solutions / Q000494:pharmacology; D010738:Type C Phospholipases / Q000378:metabolism
D003902:Detergents; D005960:Glucosides; D017261:Glycosylphosphatidylinositols; D008565:Membrane Proteins; D012996:Solutions; C018619:octyl-beta-D-glucoside; D012965:Sodium Chloride; D017830:Octoxynol; D010738:Type C Phospholipases; D006026:Glycoside Hydrolases; D043265:Phosphatidylinositol Diacylglycerol-Lyase