Proto-oncogene Pim-1: structural stability of the variants observed in tumor tissues

Pim-1 kinase belongs to the family of serine/threonine protein kinases (EC 2.7.11.1) encoded by the pim proto-oncogenes (Saris et al., 1991; Hoover et al., 1991; van der Lugt et al., 1995). Pim-1 kinase, originally identified as a common Proviral insertion site in moloney murine leukemia virus-induced T-cell lymphomas in mice (Cuypers et al., 1984), is involved in several signalling pathways and in the regulation of cell cycle progression and apoptosis. The three Pim family members Pim-1, Pim-2 and Pim-3 identified in humans have been reported as signalling protein kinases playing an important role in tumor biology (Anizon et al., 2010). Pim-1, nearly undetectable in normal tissues, is overexpressed in many haematological malignancies and in the cells of several solid tumor. In several cancer tissues Pim-1 variants have been identified and several databases for patterns of somatic mutation in human cancer genomes report mutations in this oncogene (Yuan et al., 2006; Greenman et al., 2007; Forbes et al., 2008; Akagi et al., 2009). Many of these variants are nonsynonymous single nucleotide polymorphisms (nsSNPs), single nucleotide variations occurring in the coding region and leading to amino acid substitutions (Dixit A et al., 2009). In this study we investigated the effect of amino acid substitution on the structural stability and on the activity of the Pim-1 kinase. We expressed and purified as soluble recombinant proteins some of the mutants identified in cancer and in the nsSNPs database. The mutants show a decreased thermal and thermodynamic stability and decreased activation energy relative to kinase activity, when compared to the wild- type.