Abstract
Effector triggered conformational changes of proteins such as regulators of transcription, receptors, or enzymes are the molecular basis for regulation in biology. Most proteins perform their biological functions intracellularly, in the presence of many potential interaction partners. Studies of conformational changes have mainly been performed in vitro using sophisticated physical and biochemical methods that usually require purified proteins. Here we describe the observation of conformational changes of Tet repressor in the cytoplasm of growing Escherichia coli cells, analyzed by ligand dependent disulfide crosslinking of cysteine residues substituted into mobile regions of the protein. The amount of protein undergoing the structural change is quantitatively linked to the concomitant induction of transcription of a reporter gene.
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Acknowledgements
We thank G. Unden for the E. coli strain IMW200 and E. Pook and S. Grimm for monoclonal antibodies. This work was supported by the Deutsche Forschungsgemeinschaft and the Fonds der Chemischen Industrie. B.T. was a recipient of a personal grant from the Boehringer Ingelheim Fonds.
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Tiebel, B., Garke, K. & Hillen, W. Observing conformational and activity changes of Tet repressor in vivo. Nat Struct Mol Biol 7, 479–481 (2000). https://doi.org/10.1038/nsb0600_479
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DOI: https://doi.org/10.1038/nsb0600_479
