Affinity purification and subunit structures of beta-N-acetylhexosaminidases A and B from boar epididymis.
beta-N-Acetylhexosaminidase from boar epididymis was separated into two forms, A and B, on DEAE-cellulose. Both these forms were excluded from Sepharose S-200 and had apparent Mr values of 510 000 on gradient gel electrophoresis under non-denaturing conditions. Affinity chromatography on 2-acetamido-N-(6-aminohexanoyl)-2-deoxy-beta-D-glucopyranosylam ine coupled to CNBr-activated Sepharose 4B was used to separate and purify beta-N-acetylhexosaminidases A and B that had specific activities of 115 and 380 mumol/min per mg of protein respectively. Sodium dodecyl sulphate/polyacrylamide-gel electrophoresis of denatured beta-N-acetylhexosaminidase A gave a single major component of Mr 67 000. beta-N-Acetylhexosaminidase B also had this component, and in addition had polypeptides of Mr 29 000 and 26 000. All these polypeptides were glycosylated. Antiserum to the B form precipitated form A from solution and reacted with the 67 000-Mr component or form A after electrophoretic transfer from sodium dodecyl sulphate/polyacrylamide gels to nitrocellulose sheets. The 67 000-Mr components of forms A and B yielded identical peptide maps when digested with Staphylococcus aureus V8 proteinase, and the 29 000-Mr and 26 000-Mr components in form B may be related to the 67 000-Mr polypeptide.
D000818:Animals; D002846:Chromatography, Affinity; D004591:Electrophoresis, Polyacrylamide Gel; D004822:Epididymis / Q000201:enzymology*; D006596:Hexosaminidases / Q000276:immunology / Q000302:isolation & purification; D007527:Isoenzymes / Q000302:isolation & purification; D008297:Male; D010446:Peptide Fragments / Q000032:analysis; D013552:Swine; D001619:beta-N-Acetylhexosaminidases
D007527:Isoenzymes; D010446:Peptide Fragments; D006596:Hexosaminidases; D001619:beta-N-Acetylhexosaminidases