Crystallization and preliminary X Ray analysis of Nucleoside diphosphate 1 from T. cruzi

GOMEZ BARROSO JA; MIRANDA MR; PEREIRA CA; AGUILAR CF

San Miguel de Tucumán, Tucumán

Congreso; XLV Reunión anual de la Sociedad Argentina de Investigación en Bioquímica y Biología Molecular; 2009

CRYSTALLIZATION AND PRELIMINARY X RAY ANALYSIS OF NUCLEOSIDE DIPHOSPHATE KINASE 1 FROM T. cruzi Gómez Barroso, J.A.; Miranda, M.; Pereira, C.; Aguilar, C.F. Lab. de Biol. Mol. Estructural, Univ. Nac .de San Luis, E. de los Andes 950 (5700) San Luis E-mail: jagomez@unsl.edu.ar Introduction: Trypanosoma cruzi is the etiologic agent of Chagas´ disease. The Nucleoside diphosphate kinases (NDPKs) are enzymes involved in energy management and nucleoside balance in the cell. T. cruzi TcNDPK1, a canonical isoform. The objective of this work is obtaining protein’s crystals, diffract and process the data for tridimensional structure resolution. Materials and Methods: TcNDPK1 was expressed in E. coli as a fusion protein with N-terminal His-tag. TcNDPK1 was overexpressed and purified by FPLC. Crystallization was assayed by sitting drop and hanging drop vapor diffusion method. Crystals was frozen and diffracted on synchotron x-ray radiation in Campinas (Brasil). The data set collected was reduced and merged using MOSFLM and SCALA programs. Results and Discussion: His-TcNDPK was overexpressed, purified and crystallized. The crystals are diffracted and collected the data to 3.5Å. The crystals belong to the trigonal space group P3, with unit cell parameters a=127.94, b=127.84, c=275.49. Structure determination is under way. These results will provide relevant information that could be the first step in rational drug design for treating Chagas’ disease.