Interaction of ruthenium red with Ca2+-binding proteins

doi:10.1016/0003-2697(90)90539-L

Analytical Biochemistry

Volume 188, Issue 1, July 1990, Pages 123-131

https://doi.org/10.1016/0003-2697(90)90539-L Get rights and content

Abstract

The interaction of ruthenium red, [(NH₃)₅Ru-O-Ru(NH₃)₄-O-Ru(NH₃)₅]Cl₆·4H₂O, with various Ca²⁺-binding proteins was studied. Ruthenium red inhibited Ca²⁺ binding to the sarcoplasmic reticulum protein, calsequestrin, immobilized on Sepharose 4B. Furthermore, ruthenium red bound to calsequestrin with high affinity (K_d = 0.7 μm; B_max = 218 nmol/mg protein). The dye stained calsequestrin in sodium dodecyl sulfatepolyacrylamide gels or on nitrocellulose paper and was displaced by Ca²⁺ (K_i = 1.4 mm). The specificity of ruthenium red staining of several Ca²⁺-binding proteins was investigated by comparison with two other detection methods, ⁴⁵Ca²⁺ autoradiography and the Stainsall reaction. Ruthenlum red bound to the same proteins detected by the ⁴⁵Ca²⁺ overlay technique. Ruthenium red stained both the erythrocyte Band 3 anion transporter and the Ca²⁺-ATPase of skeletal muscle sarcoplasmic reticulum. Ruthenium red also stained the EF hand conformation Ca²⁺-binding proteins, calmodulin, troponin C, and S-100. This inorganic dye provides a simple, rapid method for detecting various types of Ca²⁺-binding proteins following electrophoresis.

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Interaction of ruthenium red with Ca2+-binding proteins

Abstract

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Interaction of ruthenium red with Ca²⁺-binding proteins