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1 March 2003 Purification and Characterization of Plasma Membrane-AssociatedHuman Sperm α-l-Fucosidase
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Abstract

Detergent and salt extraction studies, as well as cytochemical localization with fluorescein isothiocyanate-bovine serum albumin-l-fucose, have provided further evidence for the plasma membrane association of a novel human sperm, α-l-fucosidase. This α-l-fucosidase has been solubilized and purified 8600-fold to high specific activity (35 000 U/mg protein) by affinity chromatography on agarose-C24-fucosylamine. To our knowledge, this is the first report concerning the purification and characterization of a mammalian plasma membrane-associated α-l-fucosidase. Both SDS-PAGE and Western blot analysis indicated the α-l-fucosidase is highly purified and contains a single subunit with a molecular mass of 51 kDa. N-glycanase studies indicated the subunit contains N-glycans, and lectin blot analysis detected the presence of mannose, but no terminal galactose or sialic acid residues. Isoelectric focusing indicated the presence of two major α-l-fucosidase isoforms (pIs 6.5 and 6.7) and a possible minor isoform (pI 6.3). Treatment of α-l-fucosidase with neuraminidase did not change its isoform profile, providing further evidence for the enzyme's lack of sialic acid residues. Kinetic analysis with 4-methylumbelliferyl α-l-fucopyranoside indicated that sperm α-l-fucosidase has a pH optimum near 7, an apparent Km of 0.08 mM, and a Vmax of 6.8 μmol/min/mg protein. The unusual properties of human sperm α-l-fucosidase argue in support of a potentially important, but presently unknown, role for this enzyme in human reproduction.

Sumpars Khunsook, Barry S. Bean, Susan R. McGowan, and Jack A. Alhadeff "Purification and Characterization of Plasma Membrane-AssociatedHuman Sperm α-l-Fucosidase," Biology of Reproduction 68(3), 709-716, (1 March 2003). https://doi.org/10.1095/biolreprod.102.004465
Received: 15 March 2002; Accepted: 1 August 2002; Published: 1 March 2003
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