Kinetic characterization of Synechocystis sp. PCC6803 1-deoxy-D-xylulose 5-phosphate reductoisomerase mutants.
Kinetic characterization of Synechocystis sp. PCC6803 1-deoxy-D-xylulose 5-phosphate reductoisomerase mutants.
复制标题
集胞藻的动力学特征。
DOI：
10.1016/j.bbapap.2005.09.003
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发表时间：
2006
期刊：
影响因子：
--
通讯作者：
Proteau,PhilipJ
中科院分区：
文献类型：
--
作者：
Fernandes,RobertaPM;Proteau,PhilipJ
甲基赤藓糖醇磷酸途径在许多细菌、植物、藻类和疟疾寄生虫恶性疟原虫中被确定为甲戊酸途径的替代途径。该途径中的第二个酶--脱氧-d-木酮糖-5-磷酸还原异构酶(DXR；E.C.1.1.1.267)自从被发现是抗菌和抗疟疾化合物磷霉素的靶标以来，一直是许多研究的焦点。大肠杆菌和运动发酵单胞菌DXR酶的一些X射线晶体结构提供了关于可能参与底物结合和催化的残基的重要结构信息。定点突变研究可以用来补充结构研究，为活性位点残基的特定变化提供动力学数据。制备了重组聚球藻的活性部位突变体。PCC6803 DXR，靶向残基D152、S153、E154、H155、M206和E223。S153和M206的改变对结合和催化作用有不同程度的影响，H155A突变对动力学参数的影响很小。
The methylerythritol phosphate pathway to isoprenoids has been firmly established as an alternate to the mevalonate pathway in many bacteria, plants, algae, and the malaria parasite Plasmodium falciparum. The second enzyme in this pathway, deoxy-d-xylulose 5-phosphate reductoisomerase (DXR; E.C. 1.1.1.267), has been the focus of many investigations since it was found to be the target of the antibacterial and antimalarial compound, fosmidomycin. Several x-ray crystal structures of the Escherichia coli and Zymomonas mobilis DXR enzymes have provided important structural information about the residues potentially involved in substrate binding and catalysis. Site-directed mutagenesis studies can be used to complement the structural studies, providing kinetic data for specific changes of active site residues. Active site mutants were prepared of the recombinant Synechocystis sp. PCC6803 DXR, targeting residues D152, S153, E154, H155, M206, and E223. Alteration of the three acidic residues had major effects on catalysis, changes to S153 and M206 had variable effects on binding and catalysis, and a H155A mutation had only minimal effects on the kinetic parameters.