Evaluating the roles of the heme a side chains in cytochrome c oxidase using designed heme proteins.
Evaluating the roles of the heme a side chains in cytochrome c oxidase using designed heme proteins.
复制标题
使用设计的血红素蛋白评估血红素 a 侧链在细胞色素 c 氧化酶中的作用。
DOI：
10.1021/bi060565t
复制
发表时间：
2006
期刊：
影响因子：
2.9
通讯作者：
Gibney,BrianR
中科院分区：
文献类型：
--
作者：
Zhuang,Jinyou;Reddi,AmitR;Wang,Zhihong;Khodaverdian,Behzad;Hegg,EricL;Gibney,BrianR
HEMEA是一种细胞色素氧化酶特有的氧化还原辅因子，对有氧呼吸至关重要。他不同于更常见的半乳糖，有两种化学修饰，即C-8甲酰基和C-2羟乙基法尼基。在设计的血红素蛋白模型上评价了这些卟啉取代基对铁和亚铁血红素结合和电化学的影响。所选的模板支架[Δ7-H3m]2是一个四α螺旋束，包含两个双(3-甲基-L-组氨酸)亚铁血红素结合位点，具有已知的绝对铁和亚铁血红素阻挡。参与血红素生物合成的hemesb、o、o+16和hemea被结合到[Δ7-H3m]2中的双(3-甲基-L-组氨酸)血红素结合部位。光谱分析表明，每个血红素中有2当量的血红素与[Δ7-H3m]2结合。血红素与模板的平衡结合研究表明，含C-2羟乙基法尼基的亚铁和亚铁形式的亚铁与亚铁的亚铁具有紧密的亲和力。结合测量的平衡中点电位，数据表明，羟乙基法呢基通过疏水相互作用使亚铁和三价铁的结合至少稳定了6.3kcal/mol。数据还表明，在血红素中加入C-8甲酰取代基导致血红素还原电位相对于血红素的179 mV或4.1千卡/摩尔的正移，这是由于铁的血红素结合相对于铁的血红素结合的不稳定。这两个取代基似乎相互抵消，以提供比亚铁和铁形式的血红素更紧密的血红素亲和力，分别至少增加6.3千卡/摩尔和2.1千卡/摩尔。这些结果也为在血红素的生物合成中观察到的反应序列提供了理论基础。
Hemeais a redox cofactor unique to cytochromecoxidases and vital to aerobic respiration. Hemeadiffers from the more common hemebby two chemical modifications, the C-8 formyl group and the C-2 hydroxyethylfarnesyl group. The effects of these porphyrin substituents on ferric and ferrous heme binding and electrochemistry were evaluated in a designed heme protein maquette. The maquette scaffold chosen, [Δ7-H3m]2, is a four-α-helix bundle that contains two bis(3-methyl-l-histidine) heme binding sites with known absolute ferric and ferrous hemebaffinities. Hemesb,o,o+16, and hemea, those involved in the biosynthesis of hemea, were incorporated into the bis(3-methyl-l-histidine) heme binding sites in [Δ7-H3m]2. Spectroscopic analyses indicate that 2 equiv of each heme binds to [Δ7-H3m]2, as designed. Equilibrium binding studies of the hemes with the maquette demonstrate the tight affinity for hemes containing the C-2 hydroxyethylfarnesyl group in both the ferric and ferrous forms. Coupled with the measured equilibrium midpoint potentials, the data indicate that the hydroxyethylfarnesyl group stabilizes the binding of both ferrous and ferric heme by at least 6.3 kcal/mol via hydrophobic interactions. The data also demonstrate that the incorporation of the C-8 formyl substituent in hemearesults in a 179 mV, or 4.1 kcal/mol, positive shift in the heme reduction potential relative to hemeodue to the destabilization of ferric heme binding relative to ferrous heme binding. The two substituents appear to counterbalance each other to provide for tighter hemeaaffinity relative to hemebin both the ferrous and ferric forms by at least 6.3 and 2.1 kcal/mol, respectively. These results also provide a rationale for the reaction sequence observed in the biosynthesis of hemea.