Abstract
A galactosyltransferase, which converts blood group O red bloodcells to B-cells, was purfied to homogeneity from plasma of blood group B subjects. The stepwise purification procedures include: (a) column chromatography with CM-Sephadex, followed by ammonium sulfate fractionation; (b) Sephadex G-200 gel filtration; (c) column chromatogr,phy with DEAE-Sephadex; and (d) column chromatography with hydroxylapatite. The procedures provided about a 400,000-fold increase of specific activity with a 40 to 50% yield. Further purification of the enzyme was performed by small scale preparative acrylamide gel electrophoresis at pH 4.3. The final enzyme preparation showed a single protein band which coincided with enzyme activity, in acrylamide gel electrophoresis, and revealed a single protein band in sodium dodecyl sulfate-gel electrophoresis. Judging from the molecular weight, which was estimated by Sephadex gel filtration, and subunit size estimated by sodium dodecyl sulfate-gel electrophoresis, the enzyme is presumably in a dimeric form. The enzyme required Mn2+ for its activity and had a pH optimum at 7.0 to 7.5.
Highlights
A galactosyltransferase, which converts blood group 0 red blood cells to B-cells, was purified to homogeneity from plasma of blood group B subjects
Further purification of the enzyme was performed by small scale preparative acrylamide gel electrophoresis at pH 4.3
The final enzyme preparation showed a single protein band which coincided with enzyme activity, in acrylamide gel electrophoresis, and revealed a single protein band in sodium dodecyl sulfate-gel electrophoresis
Summary
A galactosyltransferase, which converts blood group 0 red blood cells to B-cells, was purified to homogeneity from plasma of blood group B subjects. The stepwise purification procedures include: (a) column chromatography with CM- Sephadex G-200 gel filtration; (c) column chromatography with DEAE-Sephadex; with hydroxylapatite. Further purification of the enzyme was performed by small scale preparative acrylamide gel electrophoresis at pH 4.3.
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