The Calcium-Dependent Interaction of S100B with Its Protein Targets

Figure 5

Models for Ca²⁺-binding and then target-binding to an S100 protein. (Top) A model for the Ca²⁺-dependent interaction of S100B with target proteins involves 13 equilibrium constants (K_I to K_XIII), 11 states, and 4 conformational changes (L_I-L_IV) [109]. The most highly populated states and the predominant pathway are colored red; this is due to weak Ca²⁺ binding in the pseudo-EF-hand (site I), which greatly simplifies this model (see Scheme 1). Specifically, the binding of Ca²⁺ to the pseudo- and typical EF-hand in each S100B subunit is described by six states (A, AM_I, AM_II, AM_IM_II, BM_I, and BM_II), five equilibrium constants (K_I = [A][M]/[AM_I], K_II = [A][M]/[AM_II], K_III = [AM_I][M]/[AM_I,II], K_IV = [AM_II][M]/[AM_I,II], and K_XI = [BM_II][M]/[BM_I,II], two conformational changes (L_I: AM_II↔ BM_II, L_II: AM_I,II↔ BM_I,II) with corresponding rate constants, respectively, where A = S100B prior to the 90^∘ reorientation of helix three of S100B, B = S100B after 90^∘ reorientation of helix three, M_I = a Ca²⁺ ion bound to EF-hand I (pseudo-EF-hand), M_II = a Ca²⁺ ion bound to EF-hand II (typical EF-hand), M_I,II = Ca²⁺ ions bound to EF-hand I and EF-hand II. Upon the addition of p53 or another target (S), the model expands to 11 possible states, 13 dissociation constants, and four possible conformational changes. Whether additional equilibriums occur (K_XIV, K_XV, and K_XVI) is considered in Scheme 1. (Bottom) In a second model (Scheme 1), state A is defined as the “closed” conformation observed in the apo-state (Figure 2), and state B is after a 90^∘ reorientation of helix 3 termed the “open” conformation. In black, are states hypothesized to be populated. [A-M_II]^‡ and [B-M_II]^‡ represent short-lived intermediates, and L₁ is the Ca²⁺-dependent conformational change involving helix 3 of S100B upon binding Ca²⁺ (Figure 2). Based on NMR relaxation rate data from the PI’s lab [136], K_XIV highly favors state A. States are also considered via K_XV and K_XVI which result in B^∗ states that represent an ensemble of dynamic structures, of which, only a subset fully coordinate Ca²⁺ as observed in X-ray structures [9]. It is hypothesized that K_XV favors the B^∗M_II state(s), whereas, K_XVI favors B-M_II-S, explaining the apparent increase in Ca²⁺-binding affinity using equilibrium binding measurements that monitor free [metal ion] (Figure 3).