Dear Narayanaswamy,

You can use the same enzyme to investigate the binding of mono and divalent metals. For example in the following study the effect of metal ions (mono, divalent and heavy metals) on the activity of green crab (Scylla serrata) alkaline phosphatase was investigated. Generally the affinity of mono- and d-valent ions towards enzymes are different:

Green crab (Scylla serrata) alkaline phosphatase (EC 3.1.3.1) is a metalloenzyme, which catalyzes the nonspecific hydrolysis of phosphate monoesters. The present paper deals with the study of the effect of some kinds of metal ions on the enzyme. The positive monovalent alkali metal ions (Li(+), Na(+) and K(+)) have no effect on the enzyme; positive bivalent alkaline-earth metal ions (Mg(2+), Ca(2+) and Ba(2+)) and transition metal ions (Mn(2+), Co(2+), Ni(2+) and Cd(2+)) activate the enzyme; heavy metal ions (Hg(2+), Ag(+), Bi(2+), Cu(2+) and Zn(2+)) inhibit the enzyme. The activation of magnesium ion on the enzyme appears to be a partial noncompetitive type. The kinetic model has been set up and a new plot to determine the activation constant of Mg(2+) was put forward. From the plot, we can easily determine the activation constant (K(a)) value and the activation ratio of Mg(2+) on the enzyme. The inhibition effects of Cu(2+) and Hg(2+) on the enzyme are of noncompetitive type. The inhibition constants have been determined. The inhibition effect of Hg(2+) is stronger than that of Cu(2+).

http://www.ncbi.nlm.nih.gov/pubmed/10940645

Other examples are illustrated in the following two links:

http://www.pnas.org/content/91/15/7326.full.pdf

http://link.springer.com/article/10.1007/s10534-015-9844-x

Hoping this will be helpful,

Rafik

I have often investigated the effect of mono- and divalent cations on the activity of an enzyme using a single assay. It is important to be aware of the possibility of interference with the readout of the assay by the ions. For example, some conditions may result in precipitation of insoluble salts. Some ions may interact with a detection reagent. Some ions are colored and have an absorbance background. Some ions quench fluorescence of some fluorophores. If the assay has a coupling enzyme as part of the detection system, changing the ions in the assay may affect the activity of the coupling enzyme. You can check for such problems by making a product standard curve at each condition to see if there is any interference.

Another thing to watch for is the effect of ionic strength, as opposed to the effect of specific ions. You can make a plot of enzyme activity as a function of the ionic strength of the solution, overlaying data from all of the different salts tested, to see if this is happening. Please note that ionic strength is not the same as molarity of the salt. The definition of ionic strength is one-half times the sum of the molarities of each of the ions times the squares of their concentrations. For example, the ionic strength of ammonium sulfate is 3 times its molarity, whereas the ionic strength of sodium chloride is equal to its molarity.

i thank both the person for the answers  are very helpful thank u