It Is Widely Accepted That Certain Metal Ions Are Essential

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It is widely accepted that certain metal ions are essential to sustaining biological life. A major component in their biological relevance is that many enzymes require metal cofactors for their catalytic activity (Bertini et al., 2006). One such enzyme is Bovine Intestinal Alkaline Phosphatase or BIAP, a member of the wider enzyme classification of alkaline phosphatases. Alkaline phosphatases are dimeric metalloenzymes, enzyme proteins with metal ion cofactors directly bound to the protein, containing two Zn2+ and one Mg2+ metal-binding sites in each active site region (Kim et al., 1991). They function to catalyze the hydrolysis and transphosphorylation of phosphate monoesters. The presence of these metal ions in the enzyme structure are…show more content…
As one would expect, the deprotonation of the hydroxyl group requires a general base to accept the proton. An Mg2+ ion in coordination with a nearby water molecule can function as this general base, accepting the released proton and allowing for the deprotonation of the serine residue(Stec et al., 2000).
It is well documented that the binding of Mg2+ onto the alkaline phosphatase metal binding site is a relatively slow process when compared to other metal ions. This allows it to be outcompeted for the position by other metal ions such as Zn2+ and Co2+ (Hung et al, 2000 & Chan et al., 2002). From this ion competition, we can assume that the binding of Mg2+ is not a permanent institution; it sometimes dissociates and then reassociates with the enzyme. Therefore, basic probability tells us that if there is a higher concentration of Mg2+ ions in the enzyme solution, it will be more likely for an Mg2+ ion to be nearby to, and subsequently able to bind to, the vacated metal-binding site. Since the Mg2+ returns the enzyme to a catalytically favorable conformation, enzyme solution with a higher concentration of Mg2+ ions, should have a higher catalytic rate.
We hypothesize that the concentration of Mg2+ ions is positively correlated with the rate of hydrolysis of para-Nitrophenyl Phosphate (pNPP) by Bovine Intestinal Alkaline Phosphatase (BIAP). Our approach to test this hypothesis is to measure the absorbance of solutions after the hydrolysis of pNPP by BIAP at
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