is it possible that EDTA,Ca+ chelating agent and Ca+ have positive effect on enzyme activity?

Question

I am working with pure glucosidase enzymes, during the chractersitic study of this enzyme i found that this enzyme shows positive effect towards Ca+ and EDTA. It is quite surprising for me because EDTA is a Ca chelating agent and if EDTA added to the reaction mixture it should bind to the Ca and decrease the enzyme activity but it does not show that. Any body could tell me that is it possible that both EDTA and Ca+ positively influence the same enzyme activity. if yes then how.

I am working with Glucosidase enzyme which i purified after ion exchange column and gel filtration. I was studying the effect of metal ions on the glucosidase activity and i found that when EDTA(10mM prepared in water), added to the reaction mixture containing enzyme, and substrate (prepared in sodium citrate buffer)then it gives higher relative activity than the reaction mixture containing Ca+ under the same conditions. Is it possible, if yes then how?
Another thing is it possible that Na+ that is present in sodium citrate buffer bind to the EDTA more specifically than Ca.

Answer 1

More info is necessary.
How were the enzymes prepared and purified?
What's your exact experiment?
Do you see an increase in enzyme activity when you put both Ca and EDTA at the same time and if so at which molar ratio of the two effectors?
Do you also see activation for each effector separately? How does it compare quantitatively to the effect of the combination of the effectors?

My wild guess would be that Ca+2 is an allosteric activator, EDTA removes another metal that was co-purified with the enzyme and the latter metal is an inhibitor. If both have a positive effect on their own and an even higher when combined then it could be the case.
Another more extreme case is that EDTA removes the catalytic ion (if it can form a more stable complex than with Ca) and Ca takes its place. Then EDTA alone should kill the reaction while in combination with Ca have a positive effect. However this is a quite improbable scenario since enzymes usually have evolved for best performance and thus the enzyme would be expected to have the very common Ca+2 in the catalytic center to begin with, if Ca+2 aids the reaction more.

Answer 2

Perhaps there's a metal ion already part of the enzyme that the EDTA ligands to, separately from the Ca ions' effect?

Or maybe the EDTA is not completely encapsulating the Ca ions, and somehow increases the binding of the Ca ions to the enzyme--stabilizing or changing geometries of the 3d orbitals?

Even more likely: what form of EDTA are you using? Is it a salt? If so, perhaps the ions in the EDTA are also promoting the enzyme activity, although some Ca ions are being removed from solution.

Answer 3

Calcium can either promote or inhibit the activity of some glucosidases is some instances. I don't know which one you are working with, but the link below gives an abstract that shows the above stated effect.

If the activity of your glucosidase enzyme is calcium concentration dependent, then the reduction of free calcium by EDTA chelation may increase enzymatic activity.

Answer 4

EDTA is not a Ca2+ chelator.... It works really well for Mg2+... if you want a Ca chelator, theres another one, i cant remember it off the top of my head...

What happens when you add the EDTA to the assay with Ca? What is in your buffer? More than likely, Ca is required in your active site, the EDTA is probably binding to trace levels of Mg which is acting as an inhibitor

Answer 5

Phi D, are you thinking of ammoniumpyrrolydine diothiocarbamate, APDC? thats a good chelating agent for Ca.

Answer 6

It' POSSIBLE. The bones are where new cells are formed.