Department of Agrobioscience, Graduate School of Agriculture, Kobe University , Nada, Kobe 657-8501, Japan.
Biochemistry. 2013 Dec 17;52(50):9001-8. doi: 10.1021/bi401033y. Epub 2013 Dec 6.
Thermochromatium tepidum grows at the highest temperature among purple bacteria, and the light-harvesting 1 reaction center (LH1-RC) complex enhances the thermal stability by utilizing Ca(2+), although the molecular mechanism has yet to be resolved. Here, we applied perfusion-induced attenuated total reflection (ATR) Fourier transform infrared (FTIR) spectroscopy to highly purified LH1-RC complexes from Tch. tepidum and detected for the first time metal-sensitive fine structural changes involved in the enhanced thermal stability of this complex. The Tch. tepidum LH1-RC complex exhibited Sr(2+)/Ca(2+) ATR-FTIR difference bands that reflect changes in the polypeptide backbones and amino acid residues upon the replacement of native Ca(2+) with Sr(2+). The difference bands also appeared in the following Ca(2+)/Sr(2+) difference spectra with almost identical intensities but inverse signs, demonstrating that the structural changes induced by the metal exchange are fully reversible. In contrast, these ATR-FTIR signals were faintly detected in the mesophilic counterpart Allochromatium vinosum . A comparative analysis using LH1 complexes lacking the RCs strongly indicated that the metal-sensitive bands originate from polypeptide backbones and amino acid residues near the putative Ca(2+)-binding site at the C-terminal region of the Tch. tepidum LH1 complexes. Structural changes induced by Sr(2+) and Ba(2+) substitutions were essentially identical. However, Cd(2+) substitution exhibited unique structural modifications, which may be responsible for the severely deteriorated thermal stability of Cd(2+)-substituted complexes. Possible assignments for the present ATR-FTIR signals and their relation with the molecular mechanism of enhancing the thermal stability of Tch. tepidum LH1-RC proteins are discussed on the basis of the recent structural information on the Ca(2+)-binding site.
嗜热着色菌（Thermochromatium tepidum）是紫色光合细菌中能够生长的最高温度的细菌，其光捕获 1 反应中心（LH1-RC）复合物通过利用 Ca(2+)来增强热稳定性，尽管其分子机制尚未解决。在这里，我们应用灌注诱导的衰减全反射（ATR）傅里叶变换红外（FTIR）光谱法对来自嗜热着色菌的高度纯化的 LH1-RC 复合物进行了研究，首次检测到了该复合物增强热稳定性所涉及的金属敏感性精细结构变化。嗜热着色菌 LH1-RC 复合物表现出 Sr(2+)/Ca(2+)ATR-FTIR 差频带，反映了在 native Ca(2+)被 Sr(2+)取代时多肽骨架和氨基酸残基的变化。这些差频带也出现在 Ca(2+)/Sr(2+)差频光谱中，其强度几乎相同但符号相反，表明金属交换诱导的结构变化是完全可逆的。相比之下，在嗜温菌 Allochromatium vinosum 中，这些 ATR-FTIR 信号则微弱检测到。使用缺乏 RC 的 LH1 复合物进行的比较分析强烈表明，金属敏感性带源自嗜热着色菌 LH1 复合物 C 末端区域假定的 Ca(2+)-结合位点附近的多肽骨架和氨基酸残基。Sr(2+)和 Ba(2+)取代诱导的结构变化基本相同。然而，Cd(2+)取代表现出独特的结构修饰，这可能是 Cd(2+)-取代复合物热稳定性严重恶化的原因。根据最近关于 Ca(2+)-结合位点的结构信息，对当前 ATR-FTIR 信号的可能分配及其与嗜热着色菌 LH1-RC 蛋白增强热稳定性的分子机制之间的关系进行了讨论。
