Specific and Non-Specific Interactions in Ultra-Weak Protein-Protein Associations Revealed by Solvent Paramagnetic Relaxation Enhancements

Helle Johansson, Malene Ringkjøbing Jensen, Henrik Gesmar, Sebastian Meier, Joachim M. Vinther, Camille Keeler, Michael E. Hodsdon, Jens J. Led

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Abstract

Weak and transient protein-protein interactions underlie numerous biological processes. However, the location of the interaction sites of the specific complexes and the effect of transient, non-specific protein-protein interactions often remain elusive. We have investigated the weak self-association of human growth hormone (hGH, KD= 0.90 ± 0.03 mM) at neutral pH by the paramagnetic relaxation enhancement (PRE) of the amide protons induced by the soluble paramagnetic relaxation agent, gadodiamide (Gd(DTPA-BMA)). Primarily, it was found that the PREs are in agreement with the general Hwang-Freed model for relaxation by translational diffusion (J. Chem. Phys. 1975, 63, 4017-4025), only if crowding effects on the diffusion in the protein solution are taken into account. Secondly, by measuring the PREs of the hGH amide protons at increasing hGH concentrations and a constant concentration of the relaxation agent, it is shown that a distinction can be made between residues that are affected only by transient, non-specific protein-protein interactions and residues that are involved in specific protein-protein associations. Thus, the PREs of the former residues increase linearly with the hGH concentration in the entire concentration range because of a reduction of the diffusion caused by the transient, non-specific protein-protein interactions, while the PREs of the latter residues increase only at the lower hGH concentrations but decrease at the higher concentrations because of specific protein-protein associations that impede the access of gadodiamide to the residues of the interaction surface. Finally, it is found that the ultra-weak aggregation of hGH involves several interaction sites that are located in patches covering a large part of the protein surface.
Original languageEnglish
JournalJournal of the American Chemical Society
ISSN0002-7863
Publication statusE-pub ahead of print - 2014
Externally publishedYes

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