Expression, Purification, and Preliminary X-ray Crystal Diffraction Analysis of the Saccharomyces cerevisiae Phosphomevalonate Kinase

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Abstract Phosphomevalonate kinase (PMK) catalyzes the phosphorylation of phosphomevalonate via the mevalonate pathway to form diphosphomevalonate, an essential isoprenoid precursor. Fungal PMK, also known as Erg8p, is an essential enzyme with no orthologs found in animals, making it an attractive target for antifungal drug development. As Erg8p has only 16–17% sequence homology with previously reported PMKs structures, its reaction mechanism remains unclear due to the lack of structural information. Here, we present preliminary crystallographic studies of Saccharomyces cerevisiae Erg8p (SceErg8p). We expressed SceErg8p in Escherichia coli, purified, and obtained the protein crystals using the sitting-drop vapor-diffusion method. The best crystal exhibited a diffraction pattern with a resolution of 1.42 Å and belonged to the orthorhombic space group P21212, with unit cell parameters a = 87.2 Å, b = 110.0 Å, c = 55.7 Å. The structure of SceErg8p was solved using the AlphaFold2-predicted model. Structural refinement and characterization are currently in progress.

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