Osman Khan T, Lou Hua Jane, Qiu Wei, Brand Verena, Edwards Aled M, Turk Benjamin E, Hui Raymond
Dept of Molecular Genetics, University of Toronto, Canada; Structural Genomics Consortium, University of Toronto, Canada.
Dept of Pharmacology, Yale University, United States.
Mol Biochem Parasitol. 2015 Jun;201(2):85-9. doi: 10.1016/j.molbiopara.2015.06.002. Epub 2015 Jun 22.
FIKKs are protein kinases with distinctive sequence motifs found exclusively in Apicomplexa. Here, we report on the biochemical characterization of Plasmodium falciparum FIKK8 (PfFIKK8) and its Cryptosporidium parvum orthologue (CpFIKK) - the only member of the family predicted to be cytosolic and conserved amongst non-Plasmodium parasites. Recombinant protein samples of both were catalytically active. We characterized their phosphorylation ability using an enzymatic assay and substrate specificities using an arrayed positional scanning peptide library. Our results show that FIKK8 targets serine, preferably with arginine in the +3 and -3 positions. Furthermore, the soluble and active FIKK constructs in our experiments contained an N-terminal extension (NTE) conserved in FIKK8 orthologues from other apicomplexan species. Based on our results, we propose that this NTE is an integral feature of the FIKK subfamily.
FIKK蛋白激酶具有独特的序列基序，仅在顶复门寄生虫中发现。在此，我们报告恶性疟原虫FIKK8（PfFIKK8）及其微小隐孢子虫直系同源物（CpFIKK）的生化特性，CpFIKK是该家族中唯一预测为胞质且在非疟原虫寄生虫中保守的成员。两者的重组蛋白样品均具有催化活性。我们使用酶促测定法表征了它们的磷酸化能力，并使用阵列位置扫描肽库表征了底物特异性。我们的结果表明，FIKK8靶向丝氨酸，在+3和-3位置最好带有精氨酸。此外，我们实验中可溶性且有活性的FIKK构建体包含一个在其他顶复门物种的FIKK8直系同源物中保守的N端延伸（NTE）。基于我们的结果，我们提出这个NTE是FIKK亚家族的一个不可或缺的特征。
