Separation of β,γ-bidentate Cr · ATP diastereomers by reverse-phase high-performance liquid chromatography

✍ Scribed by Kenneth J. Gruys; Sheldon M. Schuster

• Publisher: Elsevier Science
• Year: 1982
• Tongue: English
• Weight: 592 KB
• Volume: 125
• Category: Article
• ISSN: 0003-2697
• DOI: 10.1016/0003-2697(82)90383-9

✦ Synopsis

The separation of the four diastereomers of &y-bidentate Cr. ATP using reverse-phase HPLC techniques is described. This technique provides complete resolution of the diastereomers within 10 min and relies on the use of methanesulfonic acid (in the ionized form) as an ion-pairing agent. To identify the screw sense of these resolved isomers, the CD spectra of each isomer were done, and substrate and inhibition specificities were examined using hexokinase. The results were then correlated with the isomeric assignments made by D. Dunaway-Mariano and W. W. Cleland (1980, Biochemistry 19, 1506-1515). Further studies included the monitoring of isomer interconversion at pH 6.2 to an equilibrium concentration of all four, and specific rotation measurements of the pure isomers at pH 2.5, 23°C and 546 nm.