Expression, purification, crystallization and preliminary X-ray diffraction analysis of the apo form of Ins5 2-K from Arabidopsis thaliana

J. I. Banos-Sanz, J. Sanz-Aparicio, C. A. Brearley, B. Gonzalez

Research output: Contribution to journalArticlepeer-review


Inositol 1,3,4,5,6-pentakisphosphate 2-kinase (IP5 2-K) is a key enzyme that catalyzes the synthesis of phytic acid (IP6) from inositol 1,3,4,5,6-pentakisphos­phate (IP5) and ATP. The first structure of IP5 2-K, that from Arabidopsis thaliana, has been solved previously; it only crystallized in the presence of inositol, either the substrate IP5 or the product IP6, and failed to crystallize in its free state (without inositol). Based on structural analysis, a point mutation of IP5 2-K (W129A) has been produced in order to overcome this limitation and obtain information about protein conformational changes upon substrate binding. Here, the production and crystallization of W129A IP5 2-K in its free state and with bound nucleotide is described. These crystals differed from the native crystals and belonged to the orthorhombic space group P21212, with unit-cell parameters a = 66.00, b = 68.23, c = 105.80 Å and a = 63.06, b = 71.80, c = 100.23 Å, respectively. The crystals diffracted to resolutions of 2.22 Å (apo) and 2.05 Å (nucleotide bound) using synchrotron radiation and contained one molecule per asymmetric unit. The structures have been determined using the molecular-replacement method and refinement is being undertaken.
Original languageEnglish
Pages (from-to)701-704
Number of pages4
JournalActa Crystallographica Section F
Publication statusPublished - 2012

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