Aggregation and inhibition of rat intestinal alkaline phosphatase by high concentrations of calcium. Reversibility of the processes

Abstract

Intestinal alkaline phosphatase (IAP) is an enzyme of the brush border of the enterocyte. The activity of IAP biphasically depends on calcium. Although calcium increases IAP activity, when calcium is higher than 20 mmole/L, IAP activity decreases and the amount of an aggregated form increases. The reversibility of the effect of calcium and the aggregation process are unknown. The isoelectric point of the enzyme was higher in the presence of calcium, but was the same for the enzyme and the aggregated form. The treatment with EGTA after calcium addition did not restore the enzymatic activity but produced desaggregation of the enzyme and increase in the monomeric subunits of IAP. It is concluded that the binding of calcium does not produce important modifications on the structure of the protein, that the inhibitory effect is not reversible and that calcium could be involved in the stability of the structure of the enzyme.

Regulation of intestinal calcium absorption by luminal calcium content: Role of intestinal alkaline phosphatase

“…Moreover, previous studies demonstrated that 1,25(HO) 2 vitamin D increase the expression of IAP and that IAP activity is related to calcium absorption . We have demonstrated that IAP and Ca interaction results in changes on enzyme activity and calcium absorption . However, the relationship among calcium concentration, IAP activity, and calcium absorption demonstrated in this paper is not as a consequence of changes in 1,25(OH) 2 vitamin D, as the experiment were carried out in vitro with isolated intestinal loop and within isolated duodenal cells.…”

“…Four percent stacking and 5% separating gels were used . Electrical supply was controlled with an EPS 3500 power supply (Pharmacia Biotech, Uppsala, Sweden).…”

“…Previous studies have shown that IAP expression is controlled by 1,25(OH) 2 vitamin D and that the interaction between IAP and Ca results in changes in the activity of this enzyme .…”

Luminal calcium concentration controls intestinal calcium absorption by modification of intestinal alkaline phosphatase activity

“…In those experiments, it has been demonstrated that this enzyme binds Ca and that its molecular mass increases, probably as a consequence of molecular aggregation. With regard to aggregation, the process is reversible when Ca is removed by the addition of ethylene glycol tetraacetic acid (EGTA) (10) . The results of the experiments described in the present study confirm that %Ca decreases as Ca intake increases.…”

“…Ca binds to IAP and causes changes in its activity and molecular mass (9) . The effect of Ca appears to be the consequence of Mg replacement and seems to be important in the stability of the dimeric active structure of IAP (10) . As mentioned before, the expression of proteins involved in the mechanism of Ca absorption is regulated by vitamin D (11) , in a process involving protein synthesis (12) .…”