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Weak self‐association of human growth hormone investigated by nitrogen‐15 NMR relaxation
Author(s) -
Jensen Malene Ringkjøbing,
Kristensen Søren M.,
Keeler Camille,
Christensen Hans E. M.,
Hodsdon Michael E.,
Led Jens J.
Publication year - 2008
Publication title -
proteins: structure, function, and bioinformatics
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.699
H-Index - 191
eISSN - 1097-0134
pISSN - 0887-3585
DOI - 10.1002/prot.22039
Subject(s) - dimer , chemistry , monomer , heteronuclear molecule , amide , relaxation (psychology) , oligomer , rotational correlation time , rotational diffusion , crystallography , diffusion , heteronuclear single quantum coherence spectroscopy , chemical shift , self diffusion , analytical chemistry (journal) , stereochemistry , nuclear magnetic resonance spectroscopy , molecule , chromatography , polymer , polymer chemistry , thermodynamics , organic chemistry , psychology , self service , marketing , business , social psychology , physics
The self‐association of human growth hormone (hGH) was investigated using 15 N NMR relaxation. The investigation relies on the 15 N R 1 and R 2 relaxation rates and the heteronuclear { 1 H}‐ 15 N NOEs of the backbone amide groups at multiple protein concentrations. It is shown that the rotational correlation time of hGH in solution depends strongly on its concentration, indicating a significant degree of self‐association. The self‐association is reversible and the monomers in the aggregates are noncovalently linked. Extrapolation of the relaxation data to zero concentration predicts a correlation time of 13.4 ns and a rotational diffusion anisotropy of 1.26 for monomeric hGH, in agreement with the rotational diffusion properties estimated by hydrodynamic calculations. Moreover, the extrapolation allows characterization of the backbone dynamics of monomeric hGH without interference from self‐association phenomena, and it is found that hGH is considerably more flexible than originally thought. A concerted least‐squares analysis of the 15 N relaxations and their concentration dependence reveals that the self‐association goes beyond a simple monomer‐dimer equilibrium, and that tetramers or other multimeric states co‐exist in fast exchange with the monomeric and dimeric hGH at sub‐millimolar concentrations. Small changes in the 1 H and 15 N amide chemical shifts suggest that a region around the C‐terminus is involved in the oligomer formation. Proteins 2008. © 2008 Wiley‐Liss, Inc.