期刊
JOVE-JOURNAL OF VISUALIZED EXPERIMENTS
卷 -, 期 81, 页码 -出版社
JOURNAL OF VISUALIZED EXPERIMENTS
DOI: 10.3791/50839
关键词
Chemistry; Issue 81; Molecular Chaperones; mass spectrometers; Amino Acids; Peptides; Proteins; Enzymes; Coenzymes; Protein dynamics; conformational changes; allostery; protein folding; secondary structure; mass spectrometry
资金
- Deutsche Forschungsgemeinschaft [SFB638, MA 1278/4-1]
- Deutsche Forschungsgemeinschaft (Cluster of Excellence: CellNetworks) [EXC 81/1]
All cellular processes depend on the functionality of proteins. Although the functionality of a given protein is the direct consequence of its unique amino acid sequence, it is only realized by the folding of the polypeptide chain into a single defined three-dimensional arrangement or more commonly into an ensemble of interconverting conformations. Investigating the connection between protein conformation and its function is therefore essential for a complete understanding of how proteins are able to fulfill their great variety of tasks. One possibility to study conformational changes a protein undergoes while progressing through its functional cycle is hydrogen-H-1/H-2-exchange in combination with high-resolution mass spectrometry (HX-MS). HX-MS is a versatile and robust method that adds a new dimension to structural information obtained by e. g. crystallography. It is used to study protein folding and unfolding, binding of small molecule ligands, protein-protein interactions, conformational changes linked to enzyme catalysis, and allostery. In addition, HX-MS is often used when the amount of protein is very limited or crystallization of the protein is not feasible. Here we provide a general protocol for studying protein dynamics with HX-MS and describe as an example how to reveal the interaction interface of two proteins in a complex.
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