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利什曼原虫核苷二磷酸激酶b的制备、纯化、结晶及初步X射线衍射研究

Production, purification, crystallization and preliminary X-ray diffraction studies of the nucleoside diphosphate kinase b from Leishmania major.

作者信息

Tonoli Celisa Caldana Costa, Vieira Plinio Salmazo, Ward Richard John, Arni Raghuvir Krishnaswamy, de Oliveira Arthur Henrique Cavalcante, Murakami Mario Tyago

机构信息

Center for Structural Molecular Biology, Brazilian Association for Synchrotron Light Technology, Campinas-SP, Brazil.

出版信息

Acta Crystallogr Sect F Struct Biol Cryst Commun. 2009 Nov 1;65(Pt 11):1116-9. doi: 10.1107/S1744309109037567. Epub 2009 Oct 30.

Abstract

Nucleoside diphosphate kinases (NDKs; EC 2.7.4.6) play an essential role in the synthesis of nucleotides from intermediates in the salvage pathway in all parasitic trypanosomatids and their structural studies will be instrumental in shedding light on the biochemical machinery involved in the parasite life cycle and host-parasite interactions. In this work, NDKb from Leishmania major was overexpressed in Escherichia coli, purified to homogeneity and crystallized using the sitting-drop vapour-diffusion method. The NDK crystal diffracted to 2.2 angstrom resolution and belonged to the trigonal crystal system, with unit-cell parameters a = 114.2, c = 93.9 angstrom. Translation-function calculations yielded an unambiguous solution in the enantiomorphic space group P3(2)21.

摘要

核苷二磷酸激酶(NDKs;EC 2.7.4.6)在所有寄生锥虫的补救途径中,由中间体合成核苷酸的过程中发挥着重要作用,其结构研究将有助于揭示寄生虫生命周期和宿主-寄生虫相互作用中涉及的生化机制。在这项工作中,来自硕大利什曼原虫的NDKb在大肠杆菌中过表达,纯化至同质,并采用坐滴气相扩散法进行结晶。NDK晶体的衍射分辨率为2.2埃,属于三方晶系,晶胞参数a = 114.2,c = 93.9埃。平移函数计算在对映空间群P3(2)21中得到了明确的解。

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