Solution structure of the RNA-binding cold-shock domain of the Chlamydomonas reinhardtii NAB1 protein and insights into RNA recognition.
Solution structure of the RNA-binding cold-shock domain of the Chlamydomonas reinhardtii NAB1 protein and insights into RNA recognition.
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DOI：
10.1042/bj20150217
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发表时间：
2015-07
期刊：
影响因子：
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通讯作者：
A. Sawyer;M. Landsberg;I. Ross;O. Kruse;M. Mobli;B. Hankamer
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作者：
A. Sawyer;M. Landsberg;I. Ross;O. Kruse;M. Mobli;B. Hankamer
捕光复合物（LHC）蛋白是地球上最丰富的蛋白质之一，在光合作用中起着关键作用，包括光捕获和光保护机制。莱茵衣原体核酸结合蛋白1（NAB 1）是LHC蛋白翻译的负调节因子。其N-末端冷休克结构域（CSD）结合到与光系统II（PSII）相关的特定LHC蛋白的mRNA中发现的13-nt元件[CSD共有序列（CSDCS）]，这种相互作用调节LHC表达，因此调节PSII相关天线的大小，结构和功能。在本研究中，我们阐明了NAB 1 CSD的溶液结构，通过异相NMR测定。CSD采用特征性的五链反平行β桶折叠。加入CSDCS RNA后，观察到大量NMR化学位移扰动，主要对应于典型RNA结合区域中高度保守的β2链和β3链内的表面暴露残基，但也对应于延伸阳性表面补丁和整个RNA结合位点的β链5上的残基。额外的化学位移扰动，伴随RNA结合涉及掩埋残基，这表明转录识别伴随着构象变化。我们的研究结果表明，NAB 1协会与RNA转录本通过涉及其CSD的机制是保守的祖先相关的细菌冷休克蛋白（CSP）采用的序列特异性核酸识别机制。
Light-harvesting complex (LHC) proteins are among the most abundant proteins on Earth and play critical roles in photosynthesis, both in light capture and in photoprotective mechanisms. The Chlamydomonas reinhardtii nucleic acid-binding protein 1 (NAB1) is a negative regulator of LHC protein translation. Its N-terminal cold-shock domain (CSD) binds to a 13-nt element [CSD consensus sequence (CSDCS)] found in the mRNA of specific LHC proteins associated with Photosystem II (PSII), an interaction which regulates LHC expression and, consequently, PSII-associated antenna size, structure and function. In the present study, we elucidated the solution structure of the NAB1 CSD as determined by heteronuclear NMR. The CSD adopts a characteristic five-stranded anti parallel β-barrel fold. Upon addition of CSDCS RNA, a large number of NMR chemical shift perturbations were observed, corresponding primarily to surface-exposed residues within the highly conserved β2- and β3-strands in the canonical RNA-binding region, but also to residues on β-strand 5 extending the positive surface patch and the overall RNA-binding site. Additional chemical shift perturbations that accompanied RNA binding involved buried residues, suggesting that transcript recognition is accompanied by conformational change. Our results indicate that NAB1 associates with RNA transcripts through a mechanism involving its CSD that is conserved with mechanisms of sequence-specific nucleic acid recognition employed by ancestrally related bacterial cold-shock proteins (CSPs).