墨西哥利什曼原虫：表达；鉴定和活性评估大肠埃希菌表达的重组 CRK3。
Leishmania mexicana: expression; characterization and activity assessment of E. coli-expressed recombinant CRK3.
机构信息
Department of Parasitology, Faculty of Veterinary Medicine, University of Khartoum, Sudan.
出版信息
Eur Rev Med Pharmacol Sci. 2012 Oct;16(10):1338-45.
OBJECTIVES AND METHODS
Previous studies have shown that CRK3 protein kinase of Leishmania mexicana is a potential drug target. Therefore, the aim of this study was to provide an active protein kinase for chemical inhibitors testing. A system was developed to express and affinity-purify recombinant L. mexicana CRK3 protein from Escherichia coli.
RESULTS
Biochemical analysis has confirmed the expression of the pure kinase. The bacterial-expressed kinase was found to be inactive as a monomer. The mutated CRK3-E178 protein kinase was also found to be inactive.
CONCLUSION
This study suggests that cyclin binding and phosphorylation status are both important for reconstituting protein kinase activity. Work presented by this paper has confirmed the usefulness of the prokaryotic system for production of pure homogenous recombinant protein kinase of Leishmania parasite, though this system is unable to produce active CRK3 protein kinase
目的和方法
先前的研究表明，墨西哥利什曼原虫的 CRK3 蛋白激酶是一个潜在的药物靶点。因此，本研究的目的是提供一种活性蛋白激酶，用于化学抑制剂测试。我们开发了一种从大肠杆菌中表达和亲和纯化重组 L. mexicana CRK3 蛋白的系统。
结果
生化分析证实了纯激酶的表达。发现细菌表达的激酶作为单体是无活性的。突变的 CRK3-E178 蛋白激酶也被发现是无活性的。
结论
本研究表明细胞周期蛋白结合和磷酸化状态对于重建蛋白激酶活性都很重要。本文的研究证实了原核系统在生产纯均相重组利什曼原虫蛋白激酶方面的有用性，尽管该系统无法产生有活性的 CRK3 蛋白激酶。
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