Thermodynamically Induced Conformational Changes of the Cyanobacterial Circadian Clock Protein KaiB
Thermodynamically Induced Conformational Changes of the Cyanobacterial Circadian Clock Protein KaiB
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DOI：
10.1007/s00723-011-0228-2
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发表时间：
2011-08-01
影响因子：
1
通讯作者：
Ishiura, Masahiro
中科院分区：
文献类型：
--
作者：
Mutoh, Risa;Mino, Hiroyuki;Ishiura, Masahiro
利用定点自旋标记电子自旋共振（ESR）技术研究了蓝藻生物钟蛋白KaiB的局部环境。我们制备了五个半胱氨酸残基取代的突变体KaiB标记马来酰亚胺自旋标记（MSL）。通过比较KaiBs在不同位置（Thr 64、Lys 67、Tyr 94、Gly 98和Ala 101）携带MSL的ESR谱，鉴定了局部构象变化。MSL-T64 C和MSL-K67 C的ESR谱显示MSL的相对缓慢的运动，其特征在于在4A ℃下分别为tau = 79和59 ns。MSL-Y 94 C、MSL-G98 C和MSL-A101 C的光谱显示出相对快的运动，其特征在于在4A ℃下分别为tau = 8.0、4.1和3.1 ns。这些差异是由KaiB职位的当地环境解释的。在40 A ℃孵育24 h后，所有标记KaiB的ESR谱都发生了变化，这可以用KaiB的结构弛豫来解释。
Site-directed spin labeling electron spin resonance (ESR) was applied to investigate the local environment of the cyanobacterial circadian clock protein KaiB. We prepared five cysteine residue-substituted mutants of KaiB labeled with maleimide spin label (MSL). By comparing the ESR spectra of KaiBs carrying MSL at different positions (Thr64, Lys67, Tyr94, Gly98, and Ala101), local conformational changes were identified. The ESR spectra of MSL-T64C and MSL-K67C showed the relatively slow motion of MSL characterized by tau = 79 and 59 ns at 4A degrees C, respectively. The spectra of MSL-Y94C, MSL-G98C and MSL-A101C showed relatively fast motion characterized by tau = 8.0, 4.1 and 3.1 ns at 4A degrees C, respectively. These differences were explained by the local environments of the position in KaiB. On incubation at 40A degrees C for 24 h, all ESR spectra of the labeled KaiBs changed, which can be explained by the structural relaxation of KaiB.