Issue 7, 2014

Direct monitoring of protein–protein inhibition using nano electrospray ionization mass spectrometry

Abstract

Dissociation of the TNF-alpha trimer caused by the small-molecule inhibitor SPD304 was monitored using native ESI-MS and ion mobility spectrometry (IMS). Upon addition of the inhibitor, our data clearly indicate partial dissociation of the protein into dimers and monomers. The IMS-MS analysis shows that dimeric ions have their own characteristic drift time distributions, which are different from those of the dimer ions originating in the gas phase due to collision-induced dissociation. We show that only one equivalent of the inhibitor binds to the trimeric form. We also investigated inhibition of heterodimer formation of the survival protein Bcl-xL with the cell death-promoting regions of the proteins Bak and Bad, using the small inhibitors ABT737 and ABT263. We found that ABT737 is more potent than ABT263 in preventing the heterodimerization between Bcl-xL and the Bak and Bad derived BH3 peptides. We could also monitor the mode of binding, which in this case is competitive. These results indicate that native ESI-MS can be widely used to study the inhibition of other relevant protein–protein interactions (PPIs), and provide a good basis for further improvement and identification of small-molecule PPI inhibitors.

Graphical abstract: Direct monitoring of protein–protein inhibition using nano electrospray ionization mass spectrometry

Supplementary files

Article information

Article type
Edge Article
Submitted
07 Dec 2013
Accepted
18 Mar 2014
First published
18 Mar 2014

Chem. Sci., 2014,5, 2794-2803

Author version available

Direct monitoring of protein–protein inhibition using nano electrospray ionization mass spectrometry

D. Cubrilovic, K. Barylyuk, D. Hofmann, M. J. Walczak, M. Gräber, T. Berg, G. Wider and R. Zenobi, Chem. Sci., 2014, 5, 2794 DOI: 10.1039/C3SC53360C

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