Overexpression, purification and crystallographic analysis of a unique adenosine kinase from Mycobacterium tuberculosis.
Overexpression, purification and crystallographic analysis of a unique adenosine kinase from Mycobacterium tuberculosis.
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DOI：
10.1107/s1744309105013473
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发表时间：
2005-06
期刊：
影响因子：
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通讯作者：
Yimin Wang;Mary C. Long;S. Ranganathan;V. Escuyer;W. Parker;Rongbao Li
中科院分区：
文献类型：
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作者：
Yimin Wang;Mary C. Long;S. Ranganathan;V. Escuyer;W. Parker;Rongbao Li
结核分枝杆菌的腺苷激酶是唯一一种已被分离和鉴定的原核腺苷激酶。该酶催化腺苷磷酸化为腺苷一磷酸，并参与2-甲基腺苷的活化，2-甲基腺苷是一种已证明对M具有选择活性的化合物。结核2-甲基腺苷的作用机制可能与目前的结核病治疗不同，这种化合物（或其他腺苷类似物）可能被证明是这种疾病的一种新的治疗干预。分枝结核腺苷激酶在大肠杆菌中过表达，并纯化了酶，其活性与以前报道的相当。使用气相扩散法在腺苷存在下使蛋白质结晶。晶体衍射X射线到高分辨率，并且使用同步加速器辐射收集完整的数据集到2.2A。该晶体属于空间群P3（1）21，晶胞参数a = 70.2，c = 111.6，不对称单元中含有一个蛋白质分子。通过分子置换法得到了蛋白质的初始结构模型，该模型揭示了二聚体结构。二聚体的单体相关的双重晶体学对称性。了解M.结核病腺苷激酶与人类同源物不同，这将有助于设计更有效和选择性的抗分枝杆菌剂，这些抗分枝杆菌剂被这种酶选择性激活。
Adenosine kinase from Mycobacterium tuberculosis is the only prokaryotic adenosine kinase that has been isolated and characterized. The enzyme catalyzes the phosphorylation of adenosine to adenosine monophosphate and is involved in the activation of 2-methyladenosine, a compound that has demonstrated selective activity against M. tuberculosis. The mechanism of action of 2-methyladenosine is likely to be different from those of current tuberculosis treatments and this compound (or other adenosine analogs) may prove to be a novel therapeutic intervention for this disease. The M. tuberculosis adenosine kinase was overexpressed in Escherichia coli and the enzyme was purified with activity comparable to that reported previously. The protein was crystallized in the presence of adenosine using the vapour-diffusion method. The crystals diffracted X-rays to high resolution and a complete data set was collected to 2.2 A using synchrotron radiation. The crystal belonged to space group P3(1)21, with unit-cell parameters a = 70.2, c = 111.6 A, and contained a single protein molecule in the asymmetric unit. An initial structural model of the protein was obtained by the molecular-replacement method, which revealed a dimeric structure. The monomers of the dimer were related by twofold crystallographic symmetry. An understanding of how the M. tuberculosis adenosine kinase differs from the human homolog should aid in the design of more potent and selective antimycobacterial agents that are selectively activated by this enzyme.