Crystallization and Preliminary X-Ray Diffraction Analysis of a Mammal Inositol 1,3,4,5,6-Pentakisphosphate 2-Kinase

Inositol 1,3,4,5,6-pentakisphosphate 2-kinase (IP₅ 2-K) is an enzyme that catalyses the formation of phytic acid (IP₆) from IP₅ and ATP. In mammals, IP₆ is involved in multiple events such as DNA repair and mRNA edit and it is the precursor of inositol pyrophosphates, emerging compounds shown to have an essential role in apoptosis. In addition, IP₅ 2-K have functions in cells independently of its catalytic activity, for example in rRNA biogenesis. We pursue the structure determination of a mammal IP₅ 2-K by Protein Crystallography. For this purpose, we have designed protocols for recombinant expression and purification of Mus musculus IP₅ 2-K (mIP₅ 2-K). The recombinant protein has been expressed in two different hosts, E. coli and insect cells using the LSLt and GST fusion proteins, respectively. Both macromolecule preparations yielded crystals of similar quality. Best crystals diffracted to 4.3 Å (E. coli expression) and 4.0 Å (insect cells expression) maximum resolution. Both type of crystals belong to space group P2₁2₁2₁ with an estimated solvent content compatible with the presence of two molecules per asymmetric unit. Gel filtration experiments are in agreement with this enzyme being a monomer. Crystallographic data analysis is currently undergoing.