Crystallization and preliminary X-ray crystallographic studies of chorismate synthase from Mycobacterium tuberculosis in native form and complexed with FMN.
Santos, BB1; Dias, MBV1; Elly2, F; Basso, LA2; Santos, DS2, Azevedo Jr, WF1,2.
1IBILCE - UNESP - São José do Rio Preto;
2 PUC - Porto Alegre - Rio Grande do Sul;
Recent
years have seen increases incidence of tuberculosis in both developing
and industrialized countries due the widespread emergence of
drug-resistant strains and a deadly synergy with the human immune
deficiency virus (HIV). Thus, newer and more efficient
anti-tuberculosis drugs are needed. Targets for the development
antimicrobial agents are the enzymes responsible for synthesis of
chorismate, because they are essential for inferiors organisms, but it
is absent from mammals. The last step this pathway is realized for
chorismate synthase (CS), which catalyzes the transformation of
5-enolpyruvylshikimate 3-phosphate (EPSP) to chorismate, using FMN with
a co-enzyme. In this work, we present crystallization and preliminary
X-ray cristallographic analysis of chorismate synthase from Mycobacterium tuberculosis
(MtCS) in the your native form and for the complex of MtCS:FMN. For
crystallization of MTCS was used the hanging drop vapor diffusion and
sparse matrix methods. X-ray diffraction data sets were collect at
wavelength of 1.427Ǻ using the Synchrotron Radiation Source (Station
PCr, Laboratório Nacional de Luz Sincrotron, LNLS, Campinas, Brasil)
and data were processed using the program Mosflm and Scala. The best
crystals were obtained after two days in drops containing Hepes-Na and 4 M of NaCl. Raw X-ray diffraction data for the crystal were processed to 2.22Ǻ and 2.55Ǻ of resolution for native form and for the complex of MtCS:FMN, respectively. The crystals are hexagonal, P6422
space group, with unit cell dimensions approximately of
a=b=130.7, c=157.9Å, and a=b= 90º, c= 120º, with one molecule per
asymmetric unit. Forty images were processed for each data set. The data sets presented an Rsym
of 9% and 96% completeness, approximately. With these results we intend
to determine the crystallographic structures of chorismate synthase
from M. tuberculosis, in the native form and for the complex MtCS:FMN, which will may be used in the development of new drug against tuberculosis.
Fapesp (processes 05/50608-9, 03/12472-2 and 01/07532-0).
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