IDENTIFICATION AND PURIFICATION OF DIPHOSPHOINOSITOL PENTAKISPHOSPHATE KINASE, WHICH SYNTHESIZES THE INOSITOL PYROPHOSPHATE BIS(DIPHOSPHO)INOSITOL TETRAKISPHOSPHATE

Diphosphoinositol pentakisphosphate (PP-IP5) and bis(diphospho)inosito l tetrakisphosphate (bis-PP-IP4) were recently identified as inositol phosphates which possess pyrophosphate bonds. The molecular mechanisms that regulate the cellular levels of these compounds are not yet char acterized. To pursue this question, we have previously purified an ino sitol hexakisphosphate (IP6) kinase from rat brain supernatants [Voglm aier, S. M., et al. (1996) Proc. Natl. Acad. Sci. U.S.A. 93, 4305-4310 ]. We now report the identification and purification of another novel kinase, diphosphoinositol pentakisphosphate (PP-IP5) kinase, which use s PP-IP5 as a substrate to form bis(diphospho)inositol tetrakisphospha te (bis-PP-IP4) in soluble fractions of rat forebrain. The purified pr otein, a monomer of 56 kDa, displays high affinity (K-m = 0.7 mu M) an d selectivity for PP-IP5 as a substrate. The purified enzyme also can transfer a phosphate from bis-PP-IP4 to ADP to form ATP. This ATP synt hase activity is an indication of the high phosphoryl group transfer p otential of bis-PP-IP4 and may represent a physiological role for PP-I P5 and bis-PP-IP4.