Label-Free Microscale Thermophoresis Discriminates Sites and Affinity of Protein–Ligand Binding**
نویسندگان
چکیده
Look, no label! Microscale thermophoresis makes use of the intrinsic fluorescence of proteins to quantify the binding affinities of ligands and discriminate between binding sites. This method is suitable for studying binding interactions of very small amounts of protein in solution. The binding of ligands to iGluR membrane receptors, small-molecule inhibitorss to kinase p38, aptamers to thrombin, and Ca(2+) ions to synaptotagmin was quantified.
منابع مشابه
Application Note NT003 - p38_competitive binding assay V002
p38α is considered as the key isoform involved in modulating inflammatory response in rheumatoid arthritis and inflammatory pain and is therefore a key target for compound screening in pharmaceutical industry. In this study we demonstrate the use of Microscale Thermophoresis to perform an essentially label-free measurements of small molecule binding to protein targets by using a competitive app...
متن کاملMolecular Dynamics Simulation and Free Energy Studies on the Interaction of Salicylic Acid with Human Serum Albumin (HSA)
Human serum albumin (HSA) is the most abundant protein in the blood plasma. Molecular dynamics simulations of subdomain IIA of HSA and its complex with salicylic acid (SAL) were performed to investigate structural changes induced by the ligand binding. To estimate the binding affinity of SAL molecule to subdomains IB and IIA in HSA protein, binding free energies were calculated using the Molecu...
متن کاملITC-derived binding affinity may be biased due to titrant (nano)-aggregation. Binding of halogenated benzotriazoles to the catalytic domain of human protein kinase CK2
The binding of four bromobenzotriazoles to the catalytic subunit of human protein kinase CK2 was assessed by two complementary methods: Microscale Thermophoresis (MST) and Isothermal Titration Calorimetry (ITC). New algorithm proposed for the global analysis of MST pseudo-titration data enabled reliable determination of binding affinities for two distinct sites, a relatively strong one with the...
متن کاملProtein purification-free method of binding affinity determination by microscale thermophoresis.
Quantitative characterization of protein interactions is essential in practically any field of life sciences, particularly drug discovery. Most of currently available methods of KD determination require access to purified protein of interest, generation of which can be time-consuming and expensive. We have developed a protocol that allows for determination of binding affinity by microscale ther...
متن کاملProtein-binding assays in biological liquids using microscale thermophoresis.
Protein interactions inside the human body are expected to differ from the situation in vitro. This is crucial when investigating protein functions or developing new drugs. In this study, we present a sample-efficient, free-solution method, termed microscale thermophoresis, that is capable of analysing interactions of proteins or small molecules in biological liquids such as blood serum or cell...
متن کامل