PURIFICATION AND PROPERTIES OF HUMAN BRAIN α‐L‐FUCOSIDASE

Human brain α‐L‐fucosidase has been extracted and the soluble portion has been purified 9388‐fold with 25% yield by a two‐step affinity chromatographic procedure utilizing agarose‐epsilon‐aminocaproyl‐fucosamine. Isoelectric focusing revealed that all seven isoelectric forms of the enzyme were purified. Trace amounts of eight glycosidases, with hexosaminidase being the largest contaminant (1% by activity) were found in the purified α‐L‐fucosidase preparation. Sodium dodecyl sulfate‐polyacrylamide gel electrophoresis indicated the presence of a single subunit of molecular weight 51,000 ± 2500. The purified enzyme has a pH optimum of 4.7 with a suggested second optimum of 6.6. The apparent Michaelis constant and maximal velocity of the purified enzyme with respect to the p‐nitrophenyl substrate are 0.44 mM and 10.7 μmol/min/mg protein, respectively. Ag²⁺ and Hg²⁺ completely inactivated the enzyme at concentrations of 0.1‐0.3 mM. Antibodies made previously against purified human liver α‐L‐fucosidase cross‐reacted with the purified brain α‐L‐fucosidase and gave a single precipitin line coincident with that from purified liver α‐L‐fucosidase. From all our studies it appears that at least the soluble portion of brain α‐L‐fucosidase is identical to human liver α‐L‐fucosidase.