Collect. Czech. Chem. Commun. 1993, 58, 445-451

Purification of Glycerol Kinase by "Dye-Ligand" Chromatography and Hydrophobic Interaction Chromatography on Bead-Cellulose Derivatives

Vladimír Žúbora, Albert Breiera, Marta Horváthováa, Dagmar Hagarováa, Peter Gemeinerb and Danica Mislovičováa

a Institute of Molecular Physiology and Genetics, Slovak Academy of Sciences, 833 34 Bratislava, Slovak Republic
b Institute of Chemistry, Slovak Academy of Sciences, 842 38 Bratislava, Slovak Republic


The crude extract of cytosole enzymes was obtained from homogenized cells of Saccharomyces cerevisiae by partition. The enzyme was then isolated from the lower aqueous phase displaying higher glycerol kinase activity by dye-ligand chromatography on Cibacron Blue (CB) or Remazol Brilliant Blue R (RB)-derivatized bead-cellulose, ATP being the eluent. The specific activity of glycerol kinase rised more than 10 and 7-times after affinity dye-ligand chromatography and hydrophobic interaction chromatography, respectively. Glycerol kinase obtained by the latter method was purified by CB-bead cellulose. The final preparation maintained its enzymic activity without noticeable losses during a long-term storage at 4 °C in dark.