Solubilization of membrane-bound acid alpha-glucosidase by proteolysis.
The membrane-bound acid alpha-glucosidase was purified partially (400-fold) from human placenta by solubilization with trypsin, concanavalin A-Sepharose chromatography, Ultrogel AcA-34 gel filtration, and Sephadex G-100 affinity chromatography. Two molecular forms of the enzyme were found in the final preparation of the purified enzyme. They were identical in molecular weight with a precursor (110 kDa) and an early intermediate form (105 kDa) of this enzyme. Also direct incubation of the membrane fraction without trypsin resulted in a release mainly of the 105 kDa form, which was inhibited by N-ethylmaleimide, but not by leupeptin, pepstatin or phenylmethylsulfonylfluoride. It was concluded that the precursor of acid alpha-glucosidase is an intrinsic membrane protein, which is transported into lysosomes after solubilization by proteolysis.
D005033:Ethylmaleimide / Q000494:pharmacology; D005087:Glucan 1,4-alpha-Glucosidase / Q000302:isolation & purification*; D005959:Glucosidases / Q000302:isolation & purification*; D006801:Humans; D007163:Immunosorbent Techniques; D008970:Molecular Weight; D010920:Placenta / Q000201:enzymology; D012995:Solubility; D014357:Trypsin / Q000378:metabolism
D005959:Glucosidases; D005087:Glucan 1,4-alpha-Glucosidase; D014357:Trypsin; D005033:Ethylmaleimide