title: Cloning and characterization of three Eimeria tenella lipid phosphate phosphatases.
abstract: Although lipid phosphate phosphatases (LPPs) play an important role in cellular signaling in addition to lipid biosynthesis, little is thus far known about parasite LPPs. In this study, we characterized three Eimeria tenella cDNA clones encoding LPP named EtLPP1, EtLPP2 and EtLPP3. Key structural features previously described in LPPs, including the three conserved domains proposed as catalytic sites, a single conserved N-glycosylation site, and putative transmembrane domains were discovered in the three resulting EtLPP amino acid sequences. Expression of His6-tagged EtLPP1, -2, and -3 in HEK293 cells produced immunoreactive proteins with variable molecular sizes, suggesting the presence of multiple forms of each of the three EtLPPs. The two faster-migrating protein bands below each of the three EtLPP proteins were found to be very similar to the porcine 35-kDa LPP enzyme in their molecular size and the extent of their N-glycosylation, suggesting that the three EtLPPs are partially N-glycosylated. Kinetic analyses of the activity of the three enzymes against PA, LPA, C1P and S1P showed that Km values for each of the substrates were (in μM) 284, 46, 28, and 22 for EtLPP1; 369, 179, 237, and 52 for EtLPP2; and 355, 83, and 260 for EtLPP3. However, EtLPP3 showed negligible activity on S1P. These results confirmed that the three EtLPPs have broad substrate specificity. The results also indicated that despite structural similarities, the three EtLPPs may play distinct functions through their different models of substrate preference. Furthermore, particularly high expression levels of the three EtLPP genes were detected in the sporozoite stage of the E. tenella life cycle (p<0.001), suggesting that their encoded proteins might play an important biological function in the sporozoite stage.
authors: State Key Laboratory of Veterinary Etiological Biology, Key Laboratory of Veterinary Parasitology of Gansu Province, Lanzhou Veterinary Research Institute, Chinese Academy of Agricultural Sciences, Lanzhou, Gansu, China; Jiangsu Co-Innovation Center for Prevention and Control of Important Animal Infectious Disease, Yangzhou, Jiangsu, China.
authors: Aijiang
authors: A
authors: Guo
authors: State Key Laboratory of Veterinary Etiological Biology, Key Laboratory of Veterinary Parasitology of Gansu Province, Lanzhou Veterinary Research Institute, Chinese Academy of Agricultural Sciences, Lanzhou, Gansu, China.
authors: Jianping
authors: J
authors: Cai
authors: State Key Laboratory of Veterinary Etiological Biology, Key Laboratory of Veterinary Parasitology of Gansu Province, Lanzhou Veterinary Research Institute, Chinese Academy of Agricultural Sciences, Lanzhou, Gansu, China; Jiangsu Co-Innovation Center for Prevention and Control of Important Animal Infectious Disease, Yangzhou, Jiangsu, China.
authors: Xuenong
authors: X
authors: Luo
authors: State Key Laboratory of Veterinary Etiological Biology, Key Laboratory of Veterinary Parasitology of Gansu Province, Lanzhou Veterinary Research Institute, Chinese Academy of Agricultural Sciences, Lanzhou, Gansu, China.
authors: Shaohua
authors: S
authors: Zhang
authors: State Key Laboratory of Veterinary Etiological Biology, Key Laboratory of Veterinary Parasitology of Gansu Province, Lanzhou Veterinary Research Institute, Chinese Academy of Agricultural Sciences, Lanzhou, Gansu, China.
authors: Junling
authors: J
authors: Hou
authors: State Key Laboratory of Veterinary Etiological Biology, Key Laboratory of Veterinary Parasitology of Gansu Province, Lanzhou Veterinary Research Institute, Chinese Academy of Agricultural Sciences, Lanzhou, Gansu, China.
authors: Hui
authors: H
authors: Li
authors: State Key Laboratory of Veterinary Etiological Biology, Key Laboratory of Veterinary Parasitology of Gansu Province, Lanzhou Veterinary Research Institute, Chinese Academy of Agricultural Sciences, Lanzhou, Gansu, China.
authors: Xuepeng
authors: X
authors: Cai
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D002645:Chickens; D003001:Cloning, Molecular; D016786:Eimeria tenella / Q000201:enzymology* / Q000254:growth & development; D006031:Glycosylation; D057809:HEK293 Cells; D006639:Histidine / Q000235:genetics / Q000378:metabolism; D006801:Humans; D007700:Kinetics; D008969:Molecular Sequence Data; D009842:Oligopeptides / Q000235:genetics / Q000378:metabolism; D033661:Oocysts / Q000201:enzymology; D010711:Phosphatidate Phosphatase / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis / Q000737:chemistry / Q000302:isolation & purification; D016415:Sequence Alignment; D034101:Sporozoites / Q000201:enzymology; D013379:Substrate Specificity; D013552:Swine
chemical_list: C471213:His-His-His-His-His-His; D009842:Oligopeptides; D011993:Recombinant Fusion Proteins; D006639:Histidine; C112948:lipid phosphate phosphatase; D010711:Phosphatidate Phosphatase