Two forms (I and II) of beta-N-acetyl-D-glucosaminidasefrom cock seminal plasma and one form (III) from spermatozoa were separated by chromatofocusing. The active enzyme forms I and II had pl values of 6.6 and 6.3, respectively, while form III had two subforms with pl values of 6.3 and 6.1, as determined by polyacrylamide gel electrofocusing. The optimim pH of enzyme forms I and III ranged from 3.6 to 4.0. In contrast, form II showed one distinct maximum at pH 3.7. The Km values obtained with p-nitrophenyl-beta-N-acetyl-D-glucosaminide as substrate were 0.35, 0.28, and 0.39 mM for forms I, II, and III, respectively. It is assumed that both cock spermatozoa and cock seminal plasma contain a common, enzymatically active beta-N-acetyl-D-glucosaminidase subunit with molecular weight about 32 000 and pl 6.3.