Purification and characterization of alpha-L-fucosidase from the liver of a fucosidosis patient.

Abstract

alpha-l-Fucosidase was partially purified from the liver of a fucosidosis patient by column chromatography either on agarose-epsilon-aminohexanoylfucosamine or on concanavalin A-Sepharose, despite no apparent enzymic activity in the crude liver supernatant. Mixing studies indicated that the liver of the fucosidosis patient did not lack activators or contain inhibitors of alpha-l-fucosidase activity. The partially purified alpha-l-fucosidase from the liver of the fucosidosis patient exhibits a 4-5-fold-increased Michaelis constant for the 4-methylumbelliferyl substrate (700-750mum) and a greatly decreased thermostability at 55 degrees C compared with the normal liver enzyme. The pH-activity curve is similar to that for the normal enzyme between pH5 and 8, but quite dissimilar in the acid region (pH3.0-4.5): below pH4.5 the alpha-l-fucosidase shows no activity, whereas the normal enzyme retains considerable activity (>/=50% of maximal activity). Isoelectric focusing of the alpha-l-fucosidase revealed one major form with pI5.8 and other possible minor forms. No cross-reacting material was detected when the alpha-l-fucosidase was run in double-immunodiffusion experiments against the immunoglobulin-G fraction of anti-(alpha-l-fucosidase) antibodies, but the enzyme was immunoprecipitated by this immunoglobulin-G fraction. For at least the fucosidosis patient being studied here, all the data suggest retention of a thermolabile portion of normal alpha-l-fucosidase (with characteristic Michaelis constant and pH-activity curve) or production of a kinetically altered alpha-l-fucosidase with decreased catalytic activity but antigenic similarity to the normal enzyme.

• Authors: J A Alhadeff, G L Andrews-Smith
• Journal: The Biochemical journal (Biochem J) Vol. 187 Issue 1 Pg. 45-51 (Apr 01 1980) ISSN: 0264-6021 [Print] England
• PMID: 7406870 (Publication Type: Journal Article, Research Support, U.S. Gov't, P.H.S.)

Chemical References

• Fucose
• alpha-L-Fucosidase

Topics

• Carbohydrate Metabolism, Inborn Errors (enzymology)
• Chromatography, Affinity
• Fucose (metabolism)
• Hot Temperature
• Humans
• Hydrogen-Ion Concentration
• Isoelectric Focusing
• Kinetics
• Liver (enzymology)
• alpha-L-Fucosidase (deficiency, immunology, isolation & purification)