Structural Features of Human Inositol Phosphate Multikinase Rationalize Its Inositol Phosphate Kinase and Phosphoinositide 3-Kinase Activities

The N-lobe ATP grasp fold, along with the hinge, guides ATP to transfer its γ phosphate to InsP3 to yield InsP4, which is further phosphorylated to InsP5.

“The N-lobe ATP grasp fold, along with the hinge, guides the ATP to transfer its γ phosphate to the InsP3 (1, 4, 5) cradled in the cavity created by the IP helices to yield InsP4 (1,3,4,5; 3-kinase activity), which is further phosphorylated to InsP5 (1,3,4,5, 6; 6-kinase activity).”

The first β-strand of IPMK is not always visible in all crystal structures owing to the disordered nature of the first roughly 60 amino acids.

“The first β-strand (β0 in Figure 2B) is not always visible in all crystal structures owing to the disordered nature of the first roughly 60 amino acids [41,42], which has implications for interaction with other proteins (Table 1) as well as catalytic activity.”

The protein constructs in these crystal structures deviate from wild-type human IPMK sequence as they exclude the NLS region, replaced by a short artificial linker.

“The protein constructs in these crystal structures deviate from the wild-type human IPMK sequence as they exclude the region encompassing the NLS region (residues 263–377 in pdb id 5w2g, 279–373 in pdb id 6e7f), which is replaced by a short artificial linker, to aid the crystallization of this protein.”

Both reported structures are in close agreement with a backbone root mean square deviation of 0.48 Å.

“Both reported structures are in close agreement with a backbone root mean square deviation of 0.48 Å.”

Crystal structures of human IPMK have been deposited in the PDB in free form, bound to substrates (ADP, InsP3), or inhibitors.

“Currently, different groups have deposited crystal structures of human IPMK in the PDB database in its free form (pdb id: 5w2g, 6e7f) [41,42] bound to substrates (ADP, InsP3) [42] or inhibitors [43].”

The N-lobe and C-lobe interface form a groove-like cavity that harbors the catalytic site, facilitating transfer of phosphates from ATP to inositol head groups of PIP2 and InsP3, InsP4.

“The N-lobe and the C-lobe interface form a groove-like cavity that harbors the catalytic site, which facilitates the transfer of phosphates from ATP to inositol head groups of PIP2 and InsP3, InsP4.”

Human IPMK is a 416 amino acid-long protein comprising two pseudo-domains, an N-lobe and a C-lobe, joined by a hinge.

“The human IPMK protein is a 416 amino acid-long protein comprising two pseudo-domains—an N-lobe and a C-lobe—joined by a hinge (Figure 2A).”

Removal of the region that contains the NLS results in lower affinity for the InsP3 substrate and perturbs kinetic parameters significantly.

“Furthermore, removal of the region that contains the NLS [42] (Figure 2A,B) also results in lower affinity for the InsP3 substrate and perturbs kinetic parameters significantly.”

The disordered region of ≈60 amino acids influences ATP binding such that its absence manifests in higher catalytic activity.

“Starting with the N-lobe, the disordered region of the ≈60 amino acids influences ATP binding such that its absence manifests in higher catalytic activity.”