The Effects of Zinc on Skeletal Alkaline Phosphatase Activity and Skeletal Tissues

Zinc is essential for normal growth, however, very little is known about the mechanisms by which zinc influences growth. ALP, an ectoenzyme expressed in high levels in osteoblasts, is a zinc-containing metalloenzyme and is important in bone formation. Preliminary studies showed that low levels of zinc could increase ALP specific activity in human osteosarcoma, SaOS-2 cells. Current studies were designed to characterize this zinc effect, test possible mechanisms, and determine whether the effect is physiologically relevant. Incubation with zinc resulted in a time and dose dependent increase in ALP specific activity in the extracts and conditioned media of SaOS-2 cells, and prevented the loss of ALP activity which results from phosphate withdrawal. Divalent cations other than zinc were unable to increase ALP activity, suggesting this action is unique to zinc. Normal human bone cells and MG-63 cells showed zinc-dependent increases in ALP activity similar to that observed in SaOS-2 cells. The effect of zinc was not associated with changes in protein synthesis, collagen production, ALP mRNA levels, enzyme release or ALP immunoreactive protein level. Zinc had a biphasic dose-dependent effect phosphatase activity and a small negative effect on IGF-II synthesis. Following phosphate withdrawal, some of the ALP activity loss was reversible by zinc and part was irreversible (i.e. required new protein synthesis). Zinc increased the half-life of ALP activity in intact cells and in purified and crude ALP enzyme preparations, suggesting that zinc has a direct effect on enzyme stability. Phosphate was able to inhibit the zincchelating action of EDTA and prevent the inhibition of ALP activity. The effect of zinc to increase ALP activity was not dependent on free ionic zinc, but also occurred with albumin-bound zinc. Dietary zinc intake was positively associated with ALP activity in serum and in tibial bone of adult female mice, suggesting the zinc effect may be physiologically relevant. We conclude that zinc increases ALP activity by acting directly on the enzyme, probably by stabilizing it and we hypothesize that differential zinc dissociation from the ALP active center may explain our results