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Resurveying the Tris buffer solution: the specific interaction between tris(hydroxymethyl)aminomethane and lysozyme
Li Quan1, Dengguo Wei, Xiaolu Jiang
1Beijing National Laboratory for Molecular Sciences, The Key Laboratory of Bioorganic Chemistry and Molecular Engineering, Ministry of Education, College of Chemistry, Peking University, Beijing 100871, China.
Analytical Biochemistry
|May 2, 2008
Summary
Researchers discovered a specific interaction between tris(hydroxymethyl)aminomethane (Tris) buffer and lysozyme (LZM). This finding highlights the importance of buffer selection in biochemical studies, especially for weak ligand binding assays.
Area of Science:
- Biochemistry
- Biophysics
- Molecular Biology
Background:
- Buffer selection is critical in biochemical assays.
- Tris buffer is commonly used but its interactions with proteins are not always fully understood.
- Lysozyme (LZM) is a widely studied enzyme.
Purpose of the Study:
- To investigate the specific interaction between tris(hydroxymethyl)aminomethane (Tris) and lysozyme (LZM).
- To determine the binding affinity and mode of interaction between Tris and LZM.
- To emphasize the impact of buffer choice on biochemical experiments.
Main Methods:
- Quartz crystal microbalance (QCM) biosensor for rapid interaction screening.
- Surface plasmon resonance (SPR) system for interaction validation.
- High-performance affinity chromatography (HPAC) for quantitative binding analysis.
- Molecular docking simulations to predict binding modes and energy.
Main Results:
- A specific interaction between Tris and LZM was confirmed using QCM, SPR, and HPAC.
- The apparent dissociation constant (K(D)) for LZM and immobilized Tris was determined to be 6.7 x 10(-5)M.
- Molecular docking predicted a binding free energy of -6.34 kcal mol(-1) (K(D) of 2.3 x 10(-5)M).
- Docking revealed hydrogen bonds formed between Tris hydroxyl groups and LZM residues (Asp52, Glu35, Ala107).
Conclusions:
- Tris buffer exhibits a specific binding interaction with lysozyme.
- This interaction can influence the results of lysozyme ligand binding studies.
- Avoid using Tris buffer for lysozyme weak binder studies to ensure accurate results.
