Structural requirements for key residues and auxiliary portions of a BLUF domain
Structural requirements for key residues and auxiliary portions of a BLUF domain
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DOI：
10.1021/bi8011687
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发表时间：
2008-09-30
期刊：
影响因子：
2.9
通讯作者：
Gardner, Kevin H.
中科院分区：
文献类型：
--
作者：
Wu, Qiong;Ko, Wen-Huang;Gardner, Kevin H.
球形红细菌中的BlrB是光感受器的BLUF家族中的单结构域、基于黄素的蓝光传感器蛋白。与该家族的其他成员一致，蓝光激发诱导推定的信号传导状态，其特征在于UV-可见吸收光谱中的10 nm红移。截短的BlrB构建体的结构和光谱表征确定了该蛋白质的C-末端50个氨基酸对其结构完整性是必需的，尽管不是典型的BLUF结构域架构的一部分。突变研究支持Tyr 9、Asn 33和Gln 51对于黄素结合和BLUF结构域折叠的完整性的关键作用。在黑暗和光照条件下获得的溶液NMR光谱的比较表明非常有限的光依赖性的构象变化，除了一些有趣的残基：Trp 92，Met 94，和Ile 127。值得注意的是，Ile 127侧链经历了显著的化学位移变化，尽管它在C-末端延伸中远离（类似于15埃）黄素发色团。这些数据表明，光诱导的信号从黄素通过β折叠传播到C末端延伸的最后两个α螺旋，可能提供了一种机制来传递这种变化，以启动对蓝光的细胞反应。
BlrB in Rhodobacter sphaeroides is a single domain, flavin-based blue light sensor protein in the BLUF family of photoreceptors. Consistent with other members of this family, blue light excitation induces a putative signaling state characterized by a 10 nm red shift in the UV-visible absorbance spectrum. Structural and spectroscopic characterization of truncated BlrB constructs establishes that the C-terminal 50 amino acids of this protein are essential to its structural integrity despite not being part of the canonical BLUF domain architecture. Mutagenesis studies support the critical roles of Tyr9, Asn33, and Gln51 for flavin binding and the integrity of the BLUF domain fold. Comparison of solution NMR spectra of BlrB acquired under dark and light conditions indicates very limited light-dependent conformational changes except for a few interesting residues: Trp92, Met94, and Ile127. Notably, the Ile127 side chain experiences significant chemical shift changes despite the fact that it is far (similar to 15 angstrom) from the flavin chromophore in the C-terminal extension. These data suggest that the light-induced signal is propagated from the flavin through the beta sheet to the last two alpha helices in the C-terminal extension, potentially providing a mechanism to transmit this change to initiate a cellular response to blue light.