Abstract
Affinity chromatography is one way to measure the binding constants of a protein–ligand interaction. Here we describe a method of measuring a binding constant using Ni-NTA resin to immobilize a His-tagged enzyme and the method of frontal analysis. While other methods of immobilization are possible, using the strong affinity interaction between His-tagged proteins and Ni-NTA supports results in a fast, easy, and gentle method of immobilization. Once the affinity support is created, frontal analysis can be used to measure the binding constant between the protein and various analytes.
Original language | English (US) |
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Pages (from-to) | 423-434 |
Number of pages | 12 |
Journal | Methods in Molecular Biology |
Volume | 1129 |
DOIs | |
State | Published - 2014 |
Keywords
- Affinity chromatography
- Binding constant
- Enzyme immobilization
- Frontal analysis
ASJC Scopus subject areas
- Molecular Biology
- Genetics