Affinity chromatography without immobilized ligands; A new method for studying macromolecular interactions using high-performance liquid chromatography

✍ Scribed by Shigeru Endo; Hidechika Hayashi; Akiyoshi Wada

• Publisher: Elsevier Science
• Year: 1982
• Tongue: English
• Weight: 635 KB
• Volume: 124
• Category: Article
• ISSN: 0003-2697
• DOI: 10.1016/0003-2697(82)90054-9

✦ Synopsis

A new method for studying macromolecular interactions was devised. The principle is based on affinity chromatography with a mobile zone of affinity ligand instead of a column with immobilized ligand. In this method, the difference in migration velocities between the moving zone of affinity ligand and a sample in a conventional gel-permeation column is utilized. The fast migrating zone (A zone), which is later injected into the column, and the slow migrating zone (B zone), which is injected beforehand, interfere with each other at the passing point in the column if A and B interact such as A + B @ AB. The zone interference deforms each elution profile of A and B, because the complex AB has a migration velocity different from the others. The elution profiles in zone-interference chromatography are calculated by computer simulation in the framework of the plate theory. The binding constant is calculated from the peak shift of elution volume of B in the zone-interference chromatogram. The interaction between single-strand DNA (A zone) and ribonuclease A (B zone) was studied.