Cyclodextrins (CDs) are native host systems with inherent ability to form inclusion complexes with various molecular entities, mostly hydrophobic substances. Host cyclodextrins are accommodative to water molecules as well and contain water in the native state. For β-cyclodextrin (β-CD), there is no consensus regarding the number of bound water molecules and the location of their coordination. A number of intriguing questions remain: (1) Which localities of the host’s macrocycle are the strongest attractors for the guest water molecules? (2) What are the stabilizing factors for the water clusters in the interior of β-CD and what type of interactions between water molecules and cavity walls or between the water molecules themselves are dominating the energetics of the β-CD hydration? (3) What is the maximum number of water molecules inside the cavity of β-CD? (4) How do the thermodynamic characteristics of β-CD hydration compare with those of its smaller α-cyclodextrin (α-CD) counterpart? In this study, we address these questions by employing a combination of experimental (DSC/TG) and theoretical (DFT) approaches.

Interaction between CB[n] (n = 5–8) and biologically essential mono- and divalent metal cation.

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The most widely used native cyclodextrins are α-, β- and γ-cyclodextrins containing six, seven or eight α-d-glucopyranoside units in the ring, respectively. Although the ligation properties of these host molecules have been extensively studied, a number of questions regarding their metal binding and selectivity remain unaddressed: to what extent do the size and flexibility of the host α-, β- and γ-cyclodextrins influence their metal affinity/selectivity? Which metal is the most preferred binding partner of α-, β- and γ-cyclodextrins? How do the charge, size and preferred coordination number of the metal cation shape its interactions with the host cyclodextrin? Can the guest metal cation inflict structural alterations in the host molecule and, if so, how do these changes correlate with the metal's properties? In the present study, by employing density functional theory (DFT) calculations combined with polarizable continuum model (PCM) computations, we try to answer these questions by evaluating the thermodynamic parameters of the IA, IIA and IIIA group metal binding to α, β- and γ-cyclodextrins. We assess how the interaction between the two binding partners depends on (1) the size, valence state and preferred coordination number of the guest metal cations, (2) the size and flexibility of the host molecule, and (3) the dielectric properties of the environment. The series of group IA (Na and Rb), IIA (Mg and Sr) and IIIA (Al+ and In) metal cations have been chosen for the task as they allow us to study the effect of various metal parameters (variable charge, ionic radius and coordination number) on the strength and form of the interactions with the host cyclodextrins.

Strontium salts are used for treatment of osteoporosis and bone cancer, but their impact on calcium-mediated physiological processes remains obscure. To explore Sr2+ interference with Ca2+ binding to proteins of the EF-hand family, we studied Sr2+/Ca2+ interaction with a canonical EF-hand protein, α-parvalbumin (α-PA). Evaluation of the equilibrium metal association constants for the active Ca2+ binding sites of recombinant human α-PA (‘CD’ and ‘EF’ sites) from fluorimetric titration experiments and isothermal titration calorimetry data gave 4 × 109 M−1 and 4 × 109 M−1 for Ca2+, and 2 × 107 M−1 and 2 × 106 M−1 for Sr2+. Inactivation of the EF site by homologous substitution of the Ca2+-coordinating Glu in position 12 of the EF-loop by Gln decreased Ca2+/Sr2+ affinity of the protein by an order of magnitude, whereas the analogous inactivation of the CD site induced much deeper suppression of the Ca2+/Sr2+ affinity. These results suggest that Sr2+ and Ca2+ bind to CD/EF sites of α-PA and the Ca2+/Sr2+ binding are sequential processes with the CD site being occupied first. Spectrofluorimetric Sr2+ titration of the Ca2+-loaded α-PA revealed presence of secondary Sr2+ binding site(s) with an apparent equilibrium association constant of 4 × 105 M−1. Fourier-transform infrared spectroscopy data evidence that Ca2+/Sr2+-loaded forms of α-PA exhibit similar states of their COO− groups. Near-UV circular dichroism (CD) data show that Ca2+/Sr2+ binding to α-PA induce similar changes in symmetry of microenvironment of its Phe residues. Far-UV CD experiments reveal that Ca2+/Sr2+ binding are accompanied by nearly identical changes in secondary structure of α-PA. Meanwhile, scanning calorimetry measurements show markedly lower Sr2+-induced increase in stability of tertiary structure of α-PA, compared to the Ca2+-induced effect. Theoretical modeling using Density Functional Theory computations with Polarizable Continuum Model calculations confirms that Ca2+-binding sites of α-PA are well protected against exchange of Ca2+ for Sr2+ regardless of coordination number of Sr2+, solvent exposure or rigidity of sites. The latter appears to be a key determinant of the Ca2+/Sr2+ selectivity. Overall, despite lowered affinity of α-PA to Sr2+, the latter competes with Ca2+ for the same EF-hands and induces similar structural rearrangements. The presence of a secondary Sr2+ binding site(s) could be a factor contributing to Sr2+ impact on the functional activity of proteins.