Choi Bryan, Han Calvin, LaRochelle Jonathan R, Pitsawong Warintra, Houde Damian
Relay Therapeutics, Inc., Cambridge, Massachusetts 02139, United States.
J Am Soc Mass Spectrom. 2024 Dec 4;35(12):2785-2789. doi: 10.1021/jasms.4c00334. Epub 2024 Nov 4.
Rapid equilibrium dialysis (RED) is predominantly used for the characterization of drug absorption, distribution, metabolism, and excretion (ADME) properties in plasma and biological fluids. We describe herein improvements in the use of RED in conjunction with mass spectrometry (RED-MS) to enable robust binding affinity measurements of small molecules for recombinant proteins and complexes from a single dialysis data set. The affinities calculated from RED-MS correlated well with measurements by both surface plasmon resonance (SPR) and affinity selection mass spectrometry (AS-MS). The method was particularly useful for quantifying the binding of small molecules to large protein complexes that were not amendable by common biophysical characterization techniques. Compound pooling and integration with automated liquid handling increased assay throughput and enabled the analysis of hundreds of measurements per week. RED-MS offers a viable option for measuring compound binding in solution and may facilitate small molecule affinity optimization toward difficult-to-drug protein complexes.
快速平衡透析（RED）主要用于表征血浆和生物流体中的药物吸收、分布、代谢和排泄（ADME）特性。我们在此描述了RED与质谱联用（RED-MS）的改进方法，以便从单个透析数据集中对小分子与重组蛋白及复合物进行可靠的结合亲和力测量。通过RED-MS计算得到的亲和力与表面等离子体共振（SPR）和亲和选择质谱（AS-MS）测量结果高度相关。该方法对于量化小分子与大型蛋白质复合物的结合特别有用，而这些复合物是常用生物物理表征技术无法分析的。化合物合并以及与自动液体处理的整合提高了检测通量，每周能够分析数百次测量。RED-MS为测量溶液中的化合物结合提供了一个可行的选择，并且可能有助于针对难以成药的蛋白质复合物进行小分子亲和力优化。
