Coupling of fructose-1,6-P2 to aminated agarose by Schiff base reduction. Affinity chromatography of yeast aldolase.
Fructose-1,6-P2 was immobilized by sodium borohydride reduction of the Schiff base formed with aminated agarose (AH-Sepharose 4B). The coupling occurs with high yield (25 mumoles immobilized fructose-1,6-P2 per ml packed gel) at neutral pH and room temperature. Schiff base reduction thus provides a convenient and mild coupling prodecure for sugar phosphates preserving their labile phospho ester bonds. As exemplified by a new isolation procedure for fructose-1,6-P2 aldolase from yeast, sugar phosphates insolubilized in this manner may be used for affinity chromatography of the corresponding enzymes, provided that contaminating unspecific phosphatases are removed in a preceding fractionation step.
D055598:Chemical Phenomena; D002621:Chemistry; D002846:Chromatography, Affinity / Q000379:methods*; D005634:Fructose-Bisphosphate Aldolase / Q000302:isolation & purification*; D005636:Fructosephosphates; D012441:Saccharomyces cerevisiae / Q000201:enzymology; D012545:Schiff Bases; D012685:Sepharose
D005636:Fructosephosphates; D012545:Schiff Bases; D012685:Sepharose; D005634:Fructose-Bisphosphate Aldolase