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pubmed:abstractText |
The degradation of Ins(1,3,4,5)P4 in Dictyostelium was investigated using a mixture of [3H]Ins(1,3,4,5)P4 and [3-32P]Ins-(1,3,4,5)P4. After incubation of this mixture with a Dictyostelium homogenate the 32P/3H ratio found in the InsP3 product was reduced to 24% of the ratio in the substrate. 32P-labelled inorganic phosphate was found as well, whereas hardly any InsP2 was detected. This indicates that Ins(1,3,4,5)P4 is mainly degraded by a 3-phosphatase. The other enzyme was characterized by identification of the 32P-labelled InsP3 isomer. This isomer did not co-elute with Ins(1,3,4)P3, indicating that no 5-phosphatase was present in Dictyostelium. The 32P-labelled InsP3 could be oxidized using NaIO4. The only InsP3 isomer that has these characteristics is Ins(3,4,5)P3, indicating 1-phosphatase activity. The 1-phosphatase appeared to be dependent on MgCl2, whereas the 3-phosphatase was still active in the absence of MgCl2. An analogue of Ins(1,3,4,5)P4 with a thiophosphate substitution at the 1-position was found to be almost completely resistant to hydrolysis by the 1-phosphatase, but was degraded by the 3-phosphatase.
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