Expression profiling and in silico homology modeling of Inositol pentakisphosphate 2-kinase, a potential candidate gene for low phytate trait in soybean

Low phytate soybeans are desirable both from a nutritional and economic standpoint. Inositol 1, 3, 4, 5, 6-pentakisphosphate 2-kinase (IPK1), optimizes the metabolic flux of phytate generation in soybean and thus shows much promise as a likely candidate for pathway regulation. In the present study, the differential spatial and temporal expression profiling of GmIpk1 and its two homologs Glyma06g03310 and Glyma04g03310 were carried out in Glycine max L. var Pusa 9712 revealing the early stages of seed development to be the potential target for gene manipulation. NCBI databank was screened using BLASTp to retrieve 32 plant IPK1 sequences showing high homology to GmIPK1 and its homologs. Bio-computational tools were employed to predict the protein's properties, conserved domains, and secondary structures. Using state-of-the-art in silico physicochemical approach, the three-dimensional (3D) GmIPK1 protein model (PMD ID-PM0079931), was developed based on Arabidopsis thaliana (PDB ID: 4AQK). Superimposition of 4AQK and best model of GmIPK1 revealed that the GmIPK1 aligned well and shows a sequence identity score of 54.32% with 4AQK and a low RMSD of 0.163 nm and almost similar structural features. The modeled structure was further refined considering the stereochemical geometry, energy and packing environment between the model and the template along with validation of its intrinsic dynamics. Molecular dynamics simulation studies of GmIPK1 were carried out to obtain structural insights and to understand the interactive behavior of this enzyme with ligands ADP and IP6. The results of this study provide some fundamental knowledge on the distinct mechanistic step performed by the key residues to elucidate the structure-function relationship of GmIPK1, as an initiative towards engineering "low phytate soybean".