Synthetic Inositol Phosphate Analogs Reveal that PPIP5K2 Has a Surface-Mounted Substrate Capture Site that Is a Target for Drug Discovery

摘要

class="head no_bottom_margin" id="sec1title">IntroductionThe process of signal transduction that governs many cellular activities frequently relies upon evolutionarily conserved families of small, regulatory molecules. Among them are the diphosphoinositol polyphosphates (inositol pyrophosphates: 5-PP-InsP4, 1-PP-InsP5 [1-InsP7], 5-PP-InsP5 [5-InsP7], and 1,5-[PP]2-InsP4 [InsP8]; ), in which six to eight phosphate groups are crammed around the six-carbon inositol ring. These high-energy molecules are synthesized by two distinct classes of kinases, IP6Ks and PPIP5Ks. The IP6Ks add the 5-diphosphate group (); mammals express three IP6K isoforms (). The PPIP5Ks synthesize the 1-diphosphate (); there are two isoforms in mammals (). Interest in this field has recently been heightened by demonstrations that diphosphoinositol polyphosphates operate at the interface of cell signaling and organismic homeostasis (). Here, a dynamic balance between the activities of IP6Ks and PPIP5Ks is of particular significance. For example, the synthesis of 5-PP-InsP5 by IP6Ks inhibits the PtdIns(3,4,5)P3/PDK1/AKT/mechanistic target of rapamycin (mTOR) cascade () that controls cell growth and metabolism in response to changes in levels of nutrients, growth factors, and bioenergetic status (). This inhibitory action of 5-PP-InsP5 is reversed through its further phosphorylation by the PPIP5Ks (). There may be therapeutic value in inhibiting PPIP5K activity to elevate 5-PP-InsP5 levels and attenuate the mTOR pathway, which is hyperactivated in 70% of human tumors, contributing to the derangement of cell growth and metabolism that accompanies cancer development and progression (). We recently published proof-of-principle of the latter idea by demonstrating that AKT phosphorylation in myoblasts is inhibited when PPIP5K1 expression is “knocked-down” (). It is just such therapeutic motives that frequently drive the development of drugs that can specifically target kinases such as PPIP5Ks. Candidate molecules may be rationally designed when information on protein structure is available. To this end, we recently solved the structure of the N-terminal kinase domain of PPIP5K2 (PPIP5K2^KD) in complex with natural substrate within the catalytic site (). However, the architecture of the active site exhibits substantial geometric and electrostatic constraints that raise challenges for the design of an effective yet specific inhibitor.class="figpopup" href="/pmc/articles/PMC4085797/figure/fig1/" target="figure" rid-figpopup="fig1" rid-ob="ob-fig1">Figure 1Biosynthesis of Diphosphoinositol PhosphatesIP5K, inositol pentakisphosphate 2-kinase; IP6K, inositol hexakisphosphate 5-kinase; PPIP5K, diphosphoinositol pentakisphosphate 1-kinase.