beta-D-glucuronidase is associated with goat sperm cytoskeleton

Glycosidase activities were detected as detergent-insoluble after sequentia l extractions of goat sperm with Triton X-100. Seventy percent of total bet a -glucuronidase activity was found in the detergent-insoluble fraction. Th is portion of beta -glucuronidase was resistant to extractions in the prese nce of 1 M KCl, chaotropic agents, colchicine, or cytochalasin B, being onl y partially solubilized by 3 M KCl or DNAse I treatment. The treatment with 0.1% sodium deoxycholate was effective, releasing 73% of the enzyme activi ty. Treating the deoxycholate extract with DNAse I resulted in a change in the elution profile of beta -glucuronidase as judged by gel filtration chro matography. A polyclonal antibody was developed against pancreatic beta -gl ucuronidase, and the sperm enzyme was strongly inhibited by the IgG fractio n of this antibody. Western blot analysis showed that the same protein corr espond to both Triton-soluble and insoluble enzyme. Results demonstrate tha t beta -glucuronidase is tightly bound to the Triton X-100 resistant fracti on, suggesting that the enzyme is associated to sperm cytoskeleton. J. Exp. Zool. 289:146-152, 2001. (C) 2001 Wiley-Liss, Inc.