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Oligomerization state of S100B at nanomolar concentration determined by large‐zone analytical gel filtration chromatography
Author(s) -
Drohat Alexander C.,
Weber David J.,
Nenortas Elizabeth,
Beckett Dorothy
Publication year - 1997
Publication title -
protein science
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 3.353
H-Index - 175
eISSN - 1469-896X
pISSN - 0961-8368
DOI - 10.1002/pro.5560060721
Subject(s) - dimer , size exclusion chromatography , covalent bond , chemistry , antiparallel (mathematics) , dissociation constant , dissociation (chemistry) , protein subunit , oligomer , biophysics , crystallography , chromatography , stereochemistry , biochemistry , organic chemistry , enzyme , physics , receptor , quantum mechanics , biology , magnetic field , gene
S100B is a Ca 2+ ‐binding protein known to be a non‐covalently associated dimer, S100B(ββ), at high concentrations (0.2‐3.0 mM) under reducing conditions. The solution structure of apo‐S100B(ββ) shows that the subunits associate in an antiparallel manner to form a tightly packed hydrophobic core at the dimer interface involving six of eight helices and the C‐terminal loop (Drohat AC, Amburgey JC, Abildgaard F, Starich MR, Baldisseri D, Weber DJ. 1996. Solution structure of rat apo‐S100B(ββ) as determined by NMR spectroscopy. Biochemistry 35:11577‐11588). The C‐terminal loop, however, is also known to participate in the binding of S100B to target proteins, so its participation in the dimer interface raises questions as to the physiological relevance of dimeric S100B(ββ). Therefore, we investigated the oligomerization state of S100B at low concentrations (1‐10,000 nM) using large‐zone analytical gel filtration chromatography with 35 S‐labeled S100B. We found that S100B exists (>99%) as a non‐covalently associated dimer, S100B(ββ), at 1 nM subunit concentration (500 pM dimer) in the presence or absence of saturating levels of Ca 2+ , which implies a dissociation constant in the picomolar range or lower. These results demonstrate for the first time that in reducing environments and at physiological concentrations, S100B exists as dimeric S100B(ββ) in the presence or absence of Ca 2+ , and that the non‐covalent dimer is most likely the form of S100B presented to target proteins.