The interaction of the organotin compounds trimethyltin(IV) and tributyltin(IV) chlorides with the calcium pump from sarcoplasmic reticulum membranes was studied. It was found that the presence of calcium fully protects against the inhibitory effect of both organotin compounds. However, the apparent affinity of the protein for tributyltin chloride is two orders of magnitude higher than for trimethyltin chloride (K0.5 values of 14 µ m and 1.4 m m, respectively). Studies of intrinsic fluorescence of the Ca2+‐ATPase and enzyme phosphorylation by ATP and Pi support the hypothesis that the inhibitory properties of trialkyltin compounds are due to the inhibition of calcium binding to the high‐affinity binding sites of the Ca2+‐ATPase. This suggests that there is a specific interaction between the trialkyltin compounds and the calcium binding sites of the protein. The effect of trialkyltin compounds on Ca2+‐ATPase was also addressed by differential scanning calorimetry to assess the thermal transition of the protein denaturation, and by infrared spectroscopy in the absorption region corresponding to the amide I band (1600–1700 cm−1) to observe changes in the secondary structure of the protein. We conclude that the interaction of trialkyltin compounds with Ca2+‐ATPase reduces the affinity and cooperativity for calcium binding and, consequently, the inhibition of ATPase activity. These events are accompanied by changes in the secondary structure of the protein, including loss of α‐helix structure and a concomitant increase in protein aggregation or unfolding. The activity of trialkyltin compounds on the Ca2+‐ATPase is discussed in relation to their solubility in water and in the lipid phase. Copyright © 2012 John Wiley & Sons, Ltd.

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Organotin compounds, such as tri-n-butyltin(IV) chloride (TBT), are widespread toxicants which disrupt different functions in living organisms. TBT interacts with lipid membranes and membrane proteins. The inhibition of the calcium ATPase from sarcoplasmic reticulum membranes by TBT was studied. It was found that the ATPase inhibition could not be reverted in a large time scale; moreover, an excess of TBT over enzyme did not fully inhibit the ATPase activity; therefore, it was concluded that TBT irreversibly inhibits the enzyme, and this inhibition is accompanied by a decrease in the effective TBT concentration. The residual ATP hydrolysis activity was measured at different TBT concentrations with time, and the protective effect of different calcium concentrations on the TBT inhibition was also determined. The simplest kinetic mechanism to successfully explain all the observations and the kinetic behavior was found to be a single irreversible step of the inhibitor binding to the enzyme accompanied with a first-order inhibitor inactivation. A fluorescence study of fluorescein-5-isothiocyanate-labeled enzyme revealed that TBT binding to the enzyme entails a conformational change related to the high- to low-affinity calcium-binding state transition (E1 to E2 transition), resembling the conformational change induced by vanadate linked to the formation of E2 V complex from E1 state. A docking study allowed us to propose a binding pocket for TBT in the membrane region of E1 close to the high-affinity calcium-binding sites, as well as to define the interactions with amino acid residues interfering with calcium sites occupancy.