Mapping structural interactions using in-cell NMR spectroscopy (STINT-NMR)

• Published in: Nature methods Vol. 3; no. 2; pp. 91 - 93
• Main Authors: Shekhtman, Alexander, Burz, David S, Dutta, Kaushik, Cowburn, David
• Format: Journal Article
• Language: English
• Published: United States Nature Publishing Group 01.02.2006
• Subjects: Adaptor Proteins, Signal Transducing - chemistry; Adaptor Proteins, Signal Transducing - genetics; Adaptor Proteins, Signal Transducing - metabolism; Arabinose - pharmacology; Ataxin-3; Binding Sites; Endosomal Sorting Complexes Required for Transport; Escherichia coli - genetics; Escherichia coli - metabolism; Gene Expression - drug effects; Isopropyl Thiogalactoside - pharmacology; Measurement; Methods; Models, Molecular; Nerve Tissue Proteins - chemistry; Nerve Tissue Proteins - genetics; Nerve Tissue Proteins - metabolism; NMR; Nuclear magnetic resonance; Nuclear magnetic resonance spectroscopy; Nuclear Magnetic Resonance, Biomolecular - methods; Nuclear Proteins; Peptide Fragments - chemistry; Peptide Fragments - genetics; Peptide Fragments - metabolism; Phosphoproteins - chemistry; Phosphoproteins - genetics; Phosphoproteins - metabolism; Physiological aspects; Plasmids - genetics; Protein Binding; Protein Conformation; Protein Interaction Mapping - methods; Protein Structure, Quaternary; Protein-protein interactions; Proteins - chemistry; Proteins - genetics; Proteins - metabolism; Repressor Proteins; Spectroscopy; Transfection; Ubiquitin - chemistry; Ubiquitin - genetics; Ubiquitin - metabolism; United States
• ISSN: 1548-7091; 1548-7105
• DOI: 10.1038/nmeth851

We describe a high-throughput in-cell nuclear magnetic resonance (NMR)-based method for mapping the structural changes that accompany protein-protein interactions (STINT-NMR). The method entails sequentially expressing two (or more) proteins within a single bacterial cell in a time-controlled manner and monitoring the protein interactions using in-cell NMR spectroscopy. The resulting spectra provide a complete titration of the interaction and define structural details of the interacting surfaces at atomic resolution.