Collision Induced Unfolding Classifies Ligands Bound to the Integral Membrane Translocator Protein
| Autor: | Jian Liu, Brandon T. Ruotolo, Sarah M Fantin, Shuai Niu, Sugyan M. Dixit, Shelagh Ferguson-Miller, Kristine F Parson, Daniel A. Polasky |
|---|---|
| Rok vydání: | 2019 |
| Předmět: |
Porphyrins
Protein Conformation Dimer Rhodobacter sphaeroides Ligands 010402 general chemistry Mass spectrometry 01 natural sciences Analytical Chemistry chemistry.chemical_compound Translocator protein Point Mutation Protein Unfolding Binding Sites biology Chemistry 010401 analytical chemistry Membrane Proteins Ligand (biochemistry) 0104 chemical sciences Membrane Membrane protein Biophysics biology.protein Unfolded protein response Mass spectrum Carrier Proteins |
| Zdroj: | Analytical Chemistry. 91:15469-15476 |
| ISSN: | 1520-6882 0003-2700 |
| DOI: | 10.1021/acs.analchem.9b03208 |
| Popis: | Membrane proteins represent most current therapeutic targets, yet remain understudied due to their insolubility in aqueous solvents and generally low yields during purification and expression. Ion mobility-mass spectrometry and collision induced unfolding experiments have recently garnered attention as methods capable of directly detecting and quantifying ligand binding within a wide range of membrane protein systems. Despite prior success, ionized surfactant often creates chemical noise patterns resulting in significant challenges surrounding the study of small membrane protein-ligand complexes. Here, we present a new data analysis workflow that overcomes such chemical noise and then utilize this approach to quantify and classify ligand binding associated with the 36 kDa dimer of translocator protein (TSPO). Following our denoising protocol, we detect separate gas-phase unfolding signatures for lipid and protoporphyrin TSPO binders, molecular classes that likely interact with separate regions of the protein surface. Further, a detailed classification analysis reveals that lipid alkyl chain saturation levels can be detected within our gas-phase protein unfolding data. We combine these data and classification schemes with mass spectra acquired directly from liquid-liquid extracts to propose an identity for a previously unknown endogenous TSPO ligand. |
| Databáze: | OpenAIRE |
| Externí odkaz: |
|