Design and synthesis of lipid-coupled inositol 1,2,3,4,5,6-hexakisphosphate derivatives exhibiting high-affinity binding for the HIV-1 MA domain

Abstract

The precursor of Gag protein (Pr55(Gag)) of human immunodeficiency virus, the principal structural component required for virus assembly, is known to bind d-myo-phosphatidylinositol 4,5-bisphosphate (PIP2). The N-terminus of Pr55(Gag), the MA domain, plays a critical role in the binding of Pr55(Gag) to the plasma membrane. Herein, we designed and synthesized myo-phosphatidylinositol 2,3,4,5,6-pentakisphosphate (PIP5) derivatives comprising highly phosphorylated inositol and variously modified diacylglycerol to examine the MA-binding properties. The inositol moiety was synthesized starting with myo-inositol and assembled with a hydrophobic glycerol moiety through a phosphate linkage. The Kd value for MA-binding of the PIP5 derivative 2 (Kd = 0.25 μM) was the lowest (i.e., highest affinity) of all derivatives, i.e., 70-fold lower than the Kd for the PIP2 derivative 1 (Kd = 16.9 μM) and 100-fold lower than the Kd for IP6 (Kd = 25.7 μM), suggesting the possibility that the PIP5 derivative blocks Pr55(Gag) membrane binding by competing with PIP2 in MA-binding.

A clue to unprecedented strategy to HIV eradication: “Lock-in and apoptosis”

“…As Pr55 Gag -PIP2 binding triggers the virion budding, we considered that a small molecule that binds firmly to Pr55 Gag could antagonize PIP2 in the Pr55 Gag binding. Previously, we established a highly-sensitive in vitro assay to determine the dissociation constant ( K d ) for the binding of Pr55 Gag to phosphoinositide derivatives using surface plasmon resonance (SPR) sensor analysis 10 , 11 . We found that both negatively-charged inositol phosphates and the lipophilic acyl chains of phosphoinositide were essential for a strong interaction with Pr55 Gag 10 .…”

“…We found that both negatively-charged inositol phosphates and the lipophilic acyl chains of phosphoinositide were essential for a strong interaction with Pr55 Gag 10 . As well, fully-phosphorylated inositol IP6 (phytic acid, “PA”) bound to the MA domain approximately 10 times stronger than IP3, the inositol head of PIP2 11 . Accounting for each of these details, we had synthesized an artificial phosphoinositide, DL-HIPPO (DL- H eptanoylphosphatidyl I nositol P entakis p h o sphate), as an isomeric mixture (a 1:1 mixture of D-HIPPO and L-HIPPO) (Fig.…”

Inositol phosphates: health implications, methods of analysis, and occurrence in plant foods

“…In a T-cell line, InsP 6 was also shown to be able to suppress the replication of HIV-1 (Otake et al, 1999). Recently, Tateishi et al (2014) found InsP 6 to be able to tightly bind the matrix domain of the Pr55 Gag protein, which is a critical component for the assembly of the HIV-1 virus. Subsequently, these authors coupled InsP 6 with diacylglycerol moieties to synthesize myo-phosphatidylinositol 2,3,4,5,6-pentakisphosphate derivatives.…”

Serial femtosecond X-ray diffraction of HIV-1 Gag MA-IP6 microcrystals at ambient temperature

“…any of currently approved antiretroviral drugs [11][12][13]. Based on the structural information of the MA domain and PI(4,5)P2, we recently developed a non-natural derivative of PI(4,5)P2, named L-HIPPO, which binds to the MA domain in order to eradicate HIV [14,15]. Recent progress identifying cellular interactions of HIV-1 Gag has revealed that the MA domain of the Gag is capable of binding to inositol hexaphosphate (IP6) [14,16] ( Figure 2B & C).…”

“…Based on the structural information of the MA domain and PI(4,5)P2, we recently developed a non-natural derivative of PI(4,5)P2, named L-HIPPO, which binds to the MA domain in order to eradicate HIV [14,15]. Recent progress identifying cellular interactions of HIV-1 Gag has revealed that the MA domain of the Gag is capable of binding to inositol hexaphosphate (IP6) [14,16] ( Figure 2B & C). Understanding the dynamic nature of the structural basis of these interactions at high-resolution can provide new hypotheses for HIV therapy.…”