Dissociation of Cyclic Inositol Phosphohydrolase Activity from Annexin III

Abstract

Cyclic inositol phosphohydrolase is a phosphodiesterase that cleaves the cyclic bond of cyclic inositol monophosphate. In 1990, Ross et al. (Ross, T. S., Tait, J. F., and Majerus, P. W. (1990) Science 248, 605-607) purified this enzyme from human placenta and reported that cyclic inositol phosphohydrolase is identical to annexin III. Independent confirmation of this finding has not been provided. The relative distribution of annexin III and cyclic inositol phosphohydrolase activity in rat kidney and spleen indicated that annexin III can be dissociated from cyclic inositol phosphohydrolase activity. Rat spleen contains large quantities of annexin III, but has very little cyclic inositol phosphohydrolase activity. In contrast, rat kidney, one of the richest sources of cyclic inositol phosphohydrolase activity, possesses very little (immunohistochemistry) or no (Western blot) annexin III. Similar to cytosol of human placenta, cytosol of guinea pig kidney contains both annexin III and cyclic inositol phosphohydrolase. On SDS-gel electrophoresis, guinea pig kidney annexin III has a slightly different mobility than the human placental annexin III. Human placental annexin III co-migrates with cyclic inositol phosphohydrolase on ion exchange chromatography, while guinea pig kidney annexin III is clearly dissociated from cyclic inositol phosphohydrolase on ion exchange chromatography. Both guinea pig kidney annexin III and human placental annexin III pellet with the addition of calcium and centrifugation, while cyclic inositol phosphohydrolase activity in both of these tissues remains in the supernatant. Our studies clearly show that cyclic inositol phosphohydrolase and annexin III are two different proteins.

Can Enzymatic Activity, or Otherwise, Be Inferred from Structural Studies of Annexin III?

“…13 and might be less prone to the co-purification of the true enzyme with annexin III. A recent paper (25) showed equally clearly that the results given in Refs. 9 and 13 were due to the use of inappropriate equipment for the final purification.…”

Fetal and Placental Cyclic Inositol Phosphohydrolase in Normoxia and Hypoxia

“…The homogenate was spun at 10,000 g for 20 min, and clear supernatant used for cIPH activity. cIPH activity was performed with [ 3 H]cIP substrate as described earlier [17,18]. Ten microliters of cytosol or whole blood containing 150-1,000 Ìg protein was incubated with 10,000 dpm of cIP for 0, 30, 60 and 90 min at 37°C.…”