Study On The Binding Reaction Of Metal Ions To Protein And The Competition Of Ions With Cadmium Ion Electrode

The study of interaction between metal ions and biological macromolecules is of current interest in biology and chemistry. The binding equilibrium of cadmium ion to two important proteins -Transferrin （Tf） and Calmodulin （CaM） and ions competition have been investigated by potentiometric titrations using a Cd²⁺-selective electrode.Transferrin （Tf） is an iron-binding glycoprotein,Each Tf has two ferric ion binding sites.The main function of Tf is combing free ferric irons,controlling iron metabolism.In the normal physiological state,human serum Tf is only 30% saturated by ferric irons,so it can bind other mental ions.The existed researches on the interaction between metal ions and Tf mainly used the UV different spectroscopy.This method can only reflect the combination which could cause the changes of the protein conformation.If some metal ions also bind the non-ferric ion binding sites in Tf, UV different spectroscopy could not provide any single change. In this paper,cadmium ion selection electrode was used to detect the unbound Cd²⁺ in Cd²⁺–Tf system.Our results should be more reliable than that of UV different spectroscopy. The main results are as followings:1.The results showed that the synergistic ions can promote the combination between Tf and ions.In the absence of synergistic ions, Tf and Cd²⁺ has a binding site,the calculated equilibrium constants is : lgK1=5.51.while,in the presence of synergistic ions, Tf and Cd²⁺ has two binding sites, the calculated equilibrium constants are : lgK1=6.22,lgK2=5.23. 2.The binding is pH-dependent,the lower the pH,the less binding of Cd²⁺ to apo-Tf.Calmodulin has been found to exist in many animals and plants,and has many biological functions.Recently its transmembrane and intracellular signal transmit mechanism has been developed.CaM not bind Ca²⁺ only,many non-Ca²⁺ metal ions could combine with CaM which ionic radius is similarity with Ca²⁺（0.99 ?）.It is of importance to investigate the general metal ion binding properties of CaM.We use a cadmium ion selective electrode determination of Cd²⁺ and CaM in the binding reaction,and used cadmium ion electrode as probe to detect the binding of metal ions to CaM.Experimental study concludes that:1.There are ten metal ion binding sites in CaM per molecule.Experimental data showed that there are ten Cd²⁺ binding sites in CaM per molecule.The successive equilibrium constants are lgK1=7.76,lgK2=6.88,lgK3=6.51,lgK4=6.30,lgK5=6.11,lgK6=6.01,lgK7=5.86,lgK8=5.68,lgK9=5.52,lgK10=5.39。2.When the solution exists Ca²⁺, Ca²⁺combinate the prime binding sites priority.there are six auxiliary binding sites of CaM.3.When the pH 4.5, the calculated equilibrium constants of Cd²⁺ to CaM are: lgK1=5.13, lgK2=4.95,lgK3=4.80,lgK4=4.70.With the increase of acidity, the average coordination number of Cd²⁺-CaM system was reduced.4.We used cadmium ion electrode as probe to detec the binding of the rare earth metal ions to CaM. The competition showed that all rare earth ions could bind to sites of CaM. The binding constants of rare earth ions to CaM calculated by MATLAB program are little diversity.