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Updated: Jul 6, 2026

The Importance of Correct Protein Concentration for Kinetics and Affinity Determination in Structure-function Analysis
Published on: March 17, 2010
Quantitative analysis of ternary complex kinetics by a surface immobilization method
Xiaohan Mai1, Yang Luo2, Ruoxin Fang3
1Shanghai Engineering Research Center of Ultra-precision Optical Manufacturing, Key Laboratory of Micro and Nano Photonic Structures (Ministry of Education), College of Future Information Technology, Fudan University, Shanghai, 200433, China; Quzhou Fudan Institute, 108 Minjiang Avenue, Kecheng District, Quzhou, Zhejiang Province, China.
Abstract:
Targeted protein degradation via bifunctional molecules represents a promising therapeutic strategy, where successful degradation requires efficient ternary complex formation. This study establishes an integrated framework for reliable kinetic characterization of ternary complexes by combining theoretical modeling with experimental validation. We demonstrate that binary binding parameters can effectively guide experimental design, while proper component immobilization and the use of calibrated concentrations of pre-formed binary complexes ensure measurement accuracy. Application to the CRBN/ARV-825/BRD4 system reveals strong positive cooperativity (α = 175), which correlates with exceptional degradation potency. Our approach provides a systematic methodology for quantifying ternary complex kinetics and offers a robust platform for rational degrader optimization through kinetic-guided design.
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