title: Biophysical Characterization of the Nucleoside Diphosphate Kinase of Leishmania major and Effect of the P95S Mutation.
abstract: Nucleoside diphosphate kinases (NDK; EC 2.7.4.6) are enzymes required for maintaining intracellular levels of nucleosides triphosphates (NTP) through transfer the γ-phosphoryl group from a NTP to a NDP. The enzyme is associated with several biological functions including prevention of host ATP-mediated cytolysis during pathogenic infections. Here we present the biophysical characterization of NDK from Leishmania major and the effect of a mutation on the protein structure in solution. The structural stability was analyzed since this secreted protein may act in different microenvironments at various stages of the parasite life cycle. LmNDK and P95S mutant were subjected to denaturation with pH and guanidine. Structural transitions were monitored by circular dichroism and intrinsic fluorescence tryptophan emission. Our results showed that the LmNDK is more structurally stable than other described NDKs and that the catalytically active P95S mutant in the Kpn loop presented a decrease in protein stability, indicating the importance of this proline for maintenance of the LmNDK structure.
authors.forename: Plínio S
authors.initials: PS
authors.lastname: Vieira
authors.forename: Ana P
authors.initials: AP
authors.lastname: de Jesus Santos
authors.affiliation: Departamento de Química, Faculdade de Filosofia, Ciências e Letras de Ribeirão Preto, Universidade de São Paulo, Avenida Bandeirantes, 3900, Zip code: 14041-901, Ribeirão Preto - SP, Brazil. arthurdeoliveira@ffclrp.usp.br.
authors.forename: Arthur H C
authors.initials: AH
authors.lastname: de Oliveira
mesh_terms: D000255:Adenosine Triphosphate / Q000737:chemistry; D001703:Biophysics; D018320:Leishmania major / Q000737:chemistry / Q000201:enzymology*; D009154:Mutation / Q000235:genetics; D009701:Nucleoside-Diphosphate Kinase / Q000737:chemistry* / Q000235:genetics; D009705:Nucleosides / Q000737:chemistry* / Q000235:genetics; D010766:Phosphorylation; D011392:Proline / Q000737:chemistry; D011487:Protein Conformation; D055550:Protein Stability
chemical_list: D009705:Nucleosides; D000255:Adenosine Triphosphate; D011392:Proline; D009701:Nucleoside-Diphosphate Kinase