cw - IV - Acetylgalactosaminidase from the Limpet ,

a-N-Acetylgalaetosaminidase, devoid of P-N-acetylhexosaminidase, has been purified 860-fold from the limpet, Patella uulgata. The final preparation gives one protein band on disc gel electrophoresis, sodium dodecyl sulfate-disc gel electrophoresis, and disc gel isoelectrofocusing. By Sephadex G-ZOO filtration, the molecular weight of this enzyme is 200,000 at pH 4.2 and is 45,000 at pH 7.0. The enzyme is active at pH 4.2 but inactive at pH 7.0. These results suggest that limpet a-N-acetylgalactosaminidase exists as an active oligomer in the acidic pH and an inactive monomer in the neutral or alkaline pH. In spite of its electrophoretic purity, the final enzyme preparation contains about 0.1% j3-galactosidase and 6.7% a-galactosidase activity. Although the /3galactosidase activity in the final preparation is regarded as a contaminant, the a-galactosidase activity may be due to the intrinsic activity of the enzyme for the following reasons: both a-N-acetylgalactosaminidase and a-galactosidase activities are inactivated to the same degree upon heat and pH inactivation; both enzymes exhibit identical dissociation and association behavior as a function of pH; both activities are inhibited by galactose and N-acetylgalactosamine. The physical properties of this enzyme are: pH optimum, pH 3.8; isoelectric point, pH 5.5; K, for p-nitrophenyl a-Nacetylgalactosaminide, 0.6 mn; K, for Forssman hapten glycolipid, 0.036 mu. This enzyme liberates the N-acetylgalactosamine unit from Forssman hapten glycolipid, blood group A active glycolipid, asialo bovine submaxillary glycoprotein, and blood group A active glycoproteins.