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A Cost-Effective Immobilization Method for MBP Fusion Proteins on Microtiter Plates Using a Gelatinized
Yuri Emoto1, Ryoya Katayama2,3, Emi Hibino1,4
1Laboratory of Structural Molecular Pharmacology, Graduate School of Pharmaceutical Sciences, Nagoya University, Furocho, Chikusa-ku, Nagoya 464-8601, Aichi, Japan.
Abstract:
The detection and quantification of protein-protein interactions (PPIs) is a crucial technique that often involves the use of recombinant proteins with fusion protein tags, such as maltose-binding protein (MBP) and glutathione-S-transferase (GST). In this study, we improved the cohesive and sticky properties of gelatinized starch by supplementing it with agarose, resulting in a harder gel that could coat the bottom of a microtiter plate. The resulting gelatinized starch/agarose mixture allowed for the efficient immobilization of MBP-tagged proteins on the coated plates, enabling the use of indirect ELISA-like PPI assays. By using the enzymatic activity of GST as an indicator, we succeeded in determining the dissociation constants between MBP-tagged and GST-tagged proteins on 96-well microtiter plates and a microplate reader without any expensive specialized equipment.
