Journal
ANALYTICAL BIOCHEMISTRY
Volume 378, Issue 2, Pages 144-150Publisher
ACADEMIC PRESS INC ELSEVIER SCIENCE
DOI: 10.1016/j.ab.2008.04.006
Keywords
lysozyme; tris(hydroxymethyl)aminomethane; protein-buffer binding; quartz crystal microbalance biosensor; surface plasmon resonance; molecular docking
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An unusual phenomenon, the specific interaction between tris(hydroxymethyl)aminomethane (Tris) and lysozyme (LZM), was demonstrated for the first time by rapid screen analysis of interactions using a quartz crystal microbalance (QCM) biosensor. This phenomenon was also observed in a surface plasmon resonance (SPR) system. Further study using high-performance affinity chromatography (HPAC) confirmed this specific interaction between LZM and immobilized Tris with an apparent dissociation constant (K-D) of 6.7 x 10(-5) M. Molecular docking was carried out to identify possible modes of binding between LZM and Tris linked to a binding arm. The estimated binding free energy was -6.34 kcal mol(-1), corresponding to a K-D of 2.3 x 10(-5) M, which correlated well with the experimental value. Based on the docking model, the three hydroxyl groups of Tris form intermolecular H bonds with Asp52, Glu35, and Ala107 in LZM. This study reinforces the importance of buffer selection in quantitative biochemical investigations. For a lysozyme ligand binding study, it is better to avoid using Tris when the ligands under study are weak binders. (C) 2008 Elsevier Inc. All rights reserved.
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