Coactosin-like protein functions as a stabilizing chaperone for 5-lipoxygenase: role of tryptophan 102
Coactosin-like protein functions as a stabilizing chaperone for 5-lipoxygenase: role of tryptophan 102
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DOI：
10.1042/bj20090856
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发表时间：
2010-01-01
影响因子：
4.1
通讯作者：
Radmark, Olof
中科院分区：
文献类型：
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作者：
Esser, Julia;Rakonjac, Marija;Radmark, Olof
5-LO（5-脂氧合酶）催化促炎性LT（白三烯）生物合成的两个初始步骤，其活性受到严格调节。一种最近发现的因子CLP（辅肌动蛋白样蛋白）结合5-LO并促进LT形成。在本文中，我们报告说，CLP也稳定5-LO，并防止非营业额失活的酶在WOW。在5-LO β-夹心中的阿替班残基的突变显示5-LO-Trp（102）对于与CLP结合以及对于CLP支持5-LO活性是必需的。此外，CLP的稳定作用还依赖于CLP与5-LO之间的结合。在阻止相互作用的突变（5-LO-W102 A或CLP-K131 A）后，CLP的保护作用不存在。计算的5-LO-CLP对接模型表明，CLP可以结合到5-LO的两个结构域中的额外残基，从而可能稳定5-LO结构。为了获得活细胞中CLP和5-LO之间结合的进一步支持，确定了CLP和5-LO在单核细胞系Mono Mac 6中的亚细胞定位。在这些细胞中，只有当分化的细胞用佛波醇酯引发并用离子载体刺激时，5-LO才与核部分相关联。CLP也发现了相同的再分布模式，表明这两种蛋白质以协调的方式与细胞核相关联。本研究的结果支持CLP作为陪伴支架因子的作用，影响5-LO的稳定性和活性。
The activity of 5-LO (5-lipoxygenase), which catalyses two initial steps in the biosynthesis of pro-inflammatory LTs (leukotrienes), is strictly regulated. One recently discovered factor, CLP (coactosin-like protein), binds 5-LO and promotes LT formation. In the present paper we report that CLP also stabilizes 5-LO and prevents non-turnover inactivation of the enzyme in WOW. Mutagenesis of tryptopban residues in the 5-LO beta-sandwich showed that 5-LO-Trp(102) is essential for binding to CLP, and for CLP to support 5-LO activity. In addition, the stabilizing effect also depended on binding between CLP and 5-LO. After mutations which prevent interaction (5-LO-W102A or CLP-K131A), the protective effect of CLP was absent. A calculated 5-LO-CLP docking model indicates that CLP may bind to additional residues in both domains of 5-LO, thus possibly stabilizing the 5-LO structure. To obtain further support for binding between CLP and 5-LO in a living cell, subcellular localization of CLP and 5-LO in the monocytic cell line Mono Mac 6 was determined. In these cells, 5-LO associates with a nuclear fraction only when differentiated cells are primed with phorbol ester and stimulated with ionophore. The same pattern of redistribution was found for CLP, indicating that the two proteins associate with the nucleus in a co-ordinated fashion. The results of the present Study support a role for CLP as a chaperoning scaffold factor, influencing both the stability and the activity of 5-LO.