Successive purification of several enzymes having affinities for phosphoric groups of substrates by affinity chromatography on P-cellulose.
Glucose-6-phosphate dehydrogenase, 6-phosphogluconate dehydrogenase, glutathione reductase and pyruvate kinase of Candida utilis and baker's yeast, when in anionic form, were adsorbed on a cation exchanger, P-cellulose, due to affinities similar to those for the phosphoric groups of their respective substrates; thus, glucose-6-phosphate dehydrogenase was readily eluted by either NADP+ or NADPH, glutathione reductase by NADPH, 6-phosphogluconate dehydrogenase by 6-phosphogluconate, and pyruvate kinase by either ATP or ADP. This type of chromatography may be called "affinity-adsorption-elution chromatography"; the main principle is different from that of so-called affinity-elution chromatography. Based on these findings, a large-scale procedure suitable for successive purification of several enzymes having affinities for the phosphoric groups of their substrates was devised. As an example, glucose-6-phosphate dehydrogenase was highly purified from baker's yeast and crystallized.
D002175:Candida / Q000201:enzymology; D002482:Cellulose; D002846:Chromatography, Affinity / Q000379:methods*; D005954:Glucosephosphate Dehydrogenase / Q000302:isolation & purification*; D005980:Glutathione Reductase / Q000302:isolation & purification*; D006153:Guanosine Diphosphate / Q000378:metabolism; D009249:NADP / Q000494:pharmacology; D010710:Phosphates / Q000378:metabolism*; D010734:Phosphogluconate Dehydrogenase / Q000302:isolation & purification*; D011770:Pyruvate Kinase / Q000302:isolation & purification*; D012441:Saccharomyces cerevisiae / Q000201:enzymology
D010710:Phosphates; D006153:Guanosine Diphosphate; D009249:NADP; D002482:Cellulose; D010734:Phosphogluconate Dehydrogenase; D005954:Glucosephosphate Dehydrogenase; D005980:Glutathione Reductase; D011770:Pyruvate Kinase