title: Molecular cloning and characterization of two new isoforms of the protein kinase A catalytic subunit from the human parasite Leishmania.
abstract: Leishmania are protozoan parasites that cause extensive morbidity and mortality in humans. Genes for two new isoforms of the protein kinase A catalytic subunit (PKAC) in Leishmania, Lmpkac2a and Lmpkac2b, were cloned and characterized. The predicted open reading frames for these isoforms are 93.4% identical over 338 amino acids (aa). The conserved PK catalytic cores (subdomains I-XI) are identical, while the carboxy-terminal extensions differ by only two aa. However, LmPKAC2 shares only 62% identity over the 255 aa catalytic core region with the previously described LmPKAC1 (c-lpk2). Unlike LmPKAC1, the location of the FXXF motif at the carboxy-terminus is conserved in both LmPKAC2 isoforms; however, the aa sequence, LXXF, in isoform-2a is unusual. The leishmanial isoforms can be distinguished by their NH(2)-terminal extensions, which show minimal similarity at the primary sequence level. Structural analysis of the three enzymes based on the crystal structure of mammalian PKAs predicts that both LmPKAC2 isoforms, unlike LmPKAC1, have identical alpha-helix structures in the NH(2)-terminal extension. Lmpkac2 genes are located on chromosome 35 just downstream from the leishmanial prp8 gene. This genomic organization is conserved in two species of Leishmania and Crithidia fasciculata and allowed for the partial analysis of Cfpkac2a. Phylogenetic analysis groups the two LmPKAC2 isoforms together and separately from LmPKAC1, which is more similar to the Euglena gracilis PKAC, EPK2. These findings provide the basis for additional studies on the role of the PKA family in parasite differentiation and virulence.
authors.affiliation: Department of Parasitology, Hebrew University - Hadassah Medical School, Jerusalem 91220, Israel.
authors.forename: Michal M
authors.initials: MM
authors.lastname: Siman-Tov
authors.forename: Alasdair C
authors.initials: AC
authors.lastname: Ivens
authors.forename: Charles L
authors.initials: CL
authors.lastname: Jaffe
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D020134:Catalytic Domain; D003001:Cloning, Molecular; D017868:Cyclic AMP-Dependent Protein Kinases / Q000737:chemistry / Q000235:genetics*; D016054:DNA, Protozoan / Q000737:chemistry / Q000235:genetics; D006801:Humans; D007527:Isoenzymes / Q000235:genetics; D007891:Leishmania / Q000201:enzymology / Q000235:genetics*; D008958:Models, Molecular; D008969:Molecular Sequence Data; D010802:Phylogeny; D011487:Protein Conformation; D016415:Sequence Alignment; D017422:Sequence Analysis, DNA; D017386:Sequence Homology, Amino Acid
chemical_list: D016054:DNA, Protozoan; D007527:Isoenzymes; D017868:Cyclic AMP-Dependent Protein Kinases