Identification of two protein serine/threonine kinase genes and molecular cloning of a SNF1 type protein kinase gene from Toxoplasma gondii.

We report for the first time the identification of two protein kinase genes, in Toxoplasma gondii. Based on the conserved amino acid sequence motifs of catalytic subdomains VI(b) and IX of known protein serine/threonine kinases, degenerate oligonucleotides were synthesized and used in PCR to produce 196 and 373 bp DNAs. They encoded stretches of amino acid sequences characteristic of protein serine/threonine kinases. Dot and Southern blot analysis confirmed that these PCR products were of Toxoplasma origin. Screening of a genomic library of the organism with the 196 bp PCR product as a probe yielded 3 ToxPK1 g clones. Nucleotide sequence of two of these clones, revealed that the protein encoded, TOXPK1 resembled other carbon catabolite derepressing regulatory protein kinases. Therefore, we suggest that TOXPK1 could play a role in the interconversion of active and passive life-cycle stages of this parasite. RT-PCR studies on Toxoplasma tachyzoites' total RNA suggested that ToxPK1 gene is developmentally regulated. The 373 bp PCR product, however encoded a polypeptide that resembled the catalytic subunit of other cAMP-dependent protein kinases. Hence, this protein (TOXPK2) was considered as a product of another gene, ToxPK2.