Ac. Drohat et al., OLIGOMERIZATION STATE OF S100B AT NANOMOLAR CONCENTRATION DETERMINED BY LARGE-ZONE ANALYTICAL GEL-FILTRATION CHROMATOGRAPHY, Protein science, 6(7), 1997, pp. 1577-1582
S100B is a Ca2+-binding protein known to be a noncovalently associated
dimer, S100B(PP), at high concentrations (0.2-3.0 mM) under reducing
conditions. The solution structure of apo-S100B(PP) shows that the sub
units associate in an antiparallel manner to form a tightly packed hyd
rophobic core at the dimer interface involving six of eight helices an
d the C-terminal loop (Drohat AC, Amburgey JC, Abildgaard F, Starich M
R, Baldisseri D, Weber DJ. 1996. Solution structure of rat apo-S100B(P
P) as determined by NMR spectroscopy. Biochemistry 35:11577-11588). Th
e C-terminal loop, however, is also known to participate in the bindin
g of S100B to target proteins, so its participation in the dimer inter
face raises questions as to the physiological relevance of dimeric S10
0B(beta beta). Therefore, we investigated the oligomerization state of
S100B at low concentrations (1-10,000 nM) using large-zone analytical
gel filtration chromatography with S-35-labeled S100B. We found that
S100B exists (>99%) as a non-covalently associated dimer, S100B(PP), a
t 1 nM subunit concentration (500 pM dimer) in the presence or absence
of saturating levels of Ca2+, which implies a dissociation constant i
n the picomolar range or lower. These results demonstrate for the firs
t time that in reducing environments and at physiological concentratio
ns, S100B exists as dimeric S100B(beta beta) in the presence or absenc
e of Ca2+, and that the non-covalent dimer is most likely the form of
S100B presented to target proteins.