Purification of L-Type Pyruvate Kinase from Human Liver by Affinity Chromatography on Blue-Dextran-Sepharose Column

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Abstract
L-type pyruvate kinase (ATP: pyruvate 2-O-phosphotransferase; EC 2.7.1.40) was purified from human liver by an original method. This purification included toluene extraction, ammonium sulphate fractionation, DEAE-Sephadex batchwise, CM-Sephadex batchwise with elective elution by ATP and affinity chromatography on a Blue Dextran-Sepharose column with specific elution by fructose 1,6-diphosphate. This purification procedure allowed us to obtain 6 mg protein with a specific activity of 420 IU/mg protein, i.e. a 2,690-fold purification with an overall yield of 34%. This preparation was homogeneous as judged by immuno-diffusion, acrylamide and sodium dodecyl sulphate acrylamide-gel electrophoresis.

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