Solution Structure of Zinc- and Calcium-Bound Rat S100B as Determined by Nuclear Magnetic Resonance Spectroscopy^,

Abstract

The EF-hand calcium-binding protein S100B also binds one zinc ion per subunit with a relatively high affinity (K(d) approximately 90 nM) [Wilder et al., (2003) Biochemistry 42, 13410-13421]. In this study, the structural characterization of zinc binding to calcium-loaded S100B was examined using high-resolution NMR techniques, including structural characterization of this complex in solution at atomic resolution. As with other S100 protein structures, the quaternary structure of Zn(2+)-Ca(2+)-bound S100B was found to be dimeric with helices H1, H1', H4, and H4' forming an X-type four-helix bundle at the dimer interface. NMR data together with mutational analyses are consistent with Zn(2+) coordination arising from His-15 and His-25 of one S100B subunit and from His-85 and Glu-89 of the other subunit. The addition of Zn(2+) was also found to extend helices H4 and H4' three to four residues similar to what was previously observed with the binding of target proteins to S100B. Furthermore, a kink in helix 4 was observed in Zn(2+)-Ca(2+)-bound S100B that is not in Ca(2+)-bound S100B. These structural changes upon Zn(2+)-binding could explain the 5-fold increase in affinity that Zn(2+)-Ca(2+)-bound S100B has for peptide targets such as the TRTK peptide versus Ca(2+)-bound S100B. There are also changes in the relative positioning of the two EF-hand calcium-binding domains and the respective helices comprising these EF-hands. Changes in conformation such as these could contribute to the order of magnitude higher affinity that S100B has for calcium in the presence of Zn(2+).

Divalent Metal Ion Complexes of S100B in the Absence and Presence of Pentamidine

“…The global fold of Zn 2+ ,Ca 2+ -S100B was similar to that reported previously for Ca 2+ -S100B, 43,44 the NMR structure of Zn 2+ ,Ca 2+ -S100B, 8 and several other Ca 2+ -loaded S100 proteins. 2,5 Specifically, each subunit of Zn 2+ ,Ca 2+ -S100B contained four helices (helix 1, S1-G19; helix 2, K28-L40; helix 3, E49-D61; helix 4, D69-F88) and one small antiparallel β-sheet (strand 1, K26-K28; strand 2, D69-E67) with the dimer interface aligned as a symmetric Xtype four-helix bundle comprising helices 1, 1′ and 4, 4′, respectively (Fig.…”

“…7,8 With this in mind, the 1.88-Å-resolution X-ray crystal structure for Zn 2+ ,Ca 2+ -S100B was solved and compared to that of Ca 2+ -S100B 43,44 (Fig. 5).…”

Complex Formation between S100B Protein and the p90 Ribosomal S6 Kinase (RSK) in Malignant Melanoma Is Calcium-dependent and Inhibits Extracellular Signal-regulated Kinase (ERK)-mediated Phosphorylation of RSK

“…However, another explanation is that the Ca 2ϩ binding affinity of S100 proteins can also be increased upon binding other metals and/or their physiologically relevant protein target(s). For example, it is well established in vitro that the affinity of S100B and other S100 proteins for Ca 2ϩ is increased after Zn 2ϩ binding (48,49), redox modification of critical cysteine residues (50), and/or by target binding, although the mechanism by which Ca 2ϩ binding is increased by as much as 200-fold upon target binding to S100B is still under investigation (18,(51)(52)(53).…”

Genetic polymorphism and protein conformational plasticity in the calmodulin superfamily: Two ways to promote multifunctionality

“…The threedimensional structures of various S100 isoforms are also very similar to one another and contain a symmetrical dimer of EF-hands arranged in a compact globular fold (51,(92)(93)(94). If the overall main-chain conformation of the various NCS and S100 isoforms is so similar, how can one explain the functional differences?…”