Interactions of human alkaline phosphatase isoenzymes with triazine dyes using affinity partitioning, affinity chromatography and difference spectroscopy.
Aqueous two-phase systems consisting of dextran, polyethylene glycol and dye-liganded polyethylene glycol were employed to investigate the affinity partitioning behaviour of isoenzymes of human alkaline phosphatase. Whereas in the system without a dye ligand the partition coefficients of the isoenzymes from human intestine and placenta were identical, the isoenzyme from human liver showed a significantly lower partition coefficient under the same conditions. After addition of dye-liganded polyethylene glycol two groups of dyes possessing substantial affinities to the isoenzymes were found. One, represented by Procion Yellow HE-3G, interacts specifically with the active centre of the isoenzymes. Differences in the affinity of the isoenzymes towards the individual dye ligands are caused only by the carbohydrate content, especially by the terminal sialic acid residues. The other group of dye ligands, represented by Procion Navy MX-RB, binds obviously in a more complex fashion involving other binding sites, which are only present in alkaline phosphatase of human liver. Procion Navy MX-RB was found to function as a suitable affinity ligand for the separation of human liver alkaline phosphatase from the other isoenzymes. Differences in the primary structure of two allelic forms of human placental alkaline phosphatase [(SS) and (F)] are not recognized in aqueous two-phase systems with or without dye-liganded polyethylene glycol.
Kirchberger
J
J
Institute of Biochemistry, University of Leipzig, Germany.
Domar
U
U
Kopperschläger
G
G
Stigbrand
T
T
D000469:Alkaline Phosphatase / Q000378:metabolism*; D002846:Chromatography, Affinity; D004396:Coloring Agents / Q000378:metabolism*; D006801:Humans; D007527:Isoenzymes / Q000378:metabolism*; D008099:Liver / Q000201:enzymology; D009439:Neuraminidase / Q000378:metabolism; D013057:Spectrum Analysis; D014227:Triazines / Q000378:metabolism*
D004396:Coloring Agents; D007527:Isoenzymes; D014227:Triazines; D000469:Alkaline Phosphatase; D009439:Neuraminidase