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首页> 外文期刊>Biochimica et biophysica acta: BBA: International journal of biochemistry, biophysics and molecular biololgy. Proteins and Proteomics >Structural and functional studies of Leishmania braziliensis Hsp90
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Structural and functional studies of Leishmania braziliensis Hsp90

机译：巴西利什曼原虫Hsp90的结构和功能研究

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The ubiquitous Hsp90 is critical for protein homeostasis in the cells, stabilizing "client" proteins in a functional state. Hsp90 activity depends on its ability to bind and hydrolyze ATP, involving various conformational changes that are regulated by co-chaperones, posttranslational modifications and small molecules. Compounds like geldanamycin (GA) and radicicol inhibit the Hsp90 ATPase activity by occupying the ATP binding site, which can lead client protein to degradation and also inhibit cell growth and differentiation in protozoan parasites. Our goal was to produce the recombinant Hsp90 of Leishmania braziliensis (LbHsp90) and construct of its N-terminal (LbHsp90N) and N-domain and middle-domain (LbHsp90NM), which lacks the C-terminal dimerization domain, in order to understand how Hsp90 works in protozoa. The recombinant proteins were produced folded as attested by spectroscopy experiments. Hydrodynamic experiments revealed that LbHsp90N and LbHsp90NM behaved as elongated monomers while LbHsp90 is an elongated dimer. All proteins prevented the in vitro citrate synthase and malate dehydrogenase aggregation, attesting that they have chaperone activity, and interacted with adenosine ligands with similar dissociation constants. The LbHsp90 has low ATPase activity (kcat = 0.320 min- 1) in agreement with Hsp90 orthologs, whereas the LbHsp90NM has negligible activity, suggesting the importance of the dimeric protein for this activity. The GA interacts with LbHsp90 and with its domain constructions with different affinities and also inhibits the LbHsp90 ATPase activity with an IC50 of 0.7 ??M. All these results shed light on the LbHsp90 activity and are the first step to understanding the Hsp90 molecular chaperone system in L. braziliensis. ? 2012 Elsevier B.V. All rights reserved.
机译：普遍存在的Hsp90对于细胞中的蛋白质稳态至关重要，可将“客户”蛋白质稳定在功能状态。 Hsp90的活性取决于其结合和水解ATP的能力，涉及各种构象变化，这些构象变化受伴侣蛋白伴侣，翻译后修饰和小分子的调节。像格尔德霉素（GA）和radicicol这样的化合物通过占据ATP结合位点来抑制Hsp90 ATPase活性，这可以导致客户蛋白质降解，还可以抑制原生动物寄生虫的细胞生长和分化。我们的目标是生产巴西利什曼原虫的重组Hsp90（LbHsp90），并构建其N末端（LbHsp90N）以及缺少C末端二聚化结构域的N结构域和中间结构域（LbHsp90NM），以了解如何Hsp90在原生动物中起作用。如通过光谱学实验证明的，产生折叠的重组蛋白。流体力学实验表明，LbHsp90N和LbHsp90NM表现为细长的单体，而LbHsp90是细长的二聚体。所有蛋白均阻止了体外柠檬酸盐合酶和苹果酸脱氢酶的聚集，证明它们具有分子伴侣活性，并与具有相似解离常数的腺苷配体相互作用。与Hsp90直系同源物一致，LbHsp90具有较低的ATPase活性（kcat = 0.320 min-1），而LbHsp90NM具有微不足道的活性，表明二聚体蛋白对该活性的重要性。 GA与LbHsp90及其具有不同亲和力的结构域相互作用，并抑制LbHsp90 ATPase活性，IC50为0.7？M。所有这些结果揭示了LbHsp90的活性，是了解巴西乳杆菌Hsp90分子伴侣系统的第一步。 ？ 2012 Elsevier B.V.保留所有权利。

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