Shinzawa-Itoh Kyoko, Hatanaka Miki, Fujita Kazuya, Yano Naomine, Ogasawara Yumi, Iwata Jun, Yamashita Eiki, Tsukihara Tomitake, Yoshikawa Shinya, Muramoto Kazumasa
Graduate School of Life Science, University of Hyogo, 3-2-1 Kouto, Kamigori, Ako, Hyogo 678-1297, Japan.
School of Science, University of Hyogo, 3-2-1 Kouto, Kamigori, Ako, Hyogo 678-1297, Japan.
BBA Adv. 2021 Mar 21;1:100009. doi: 10.1016/j.bbadva.2021.100009. eCollection 2021.
Cytochrome oxidase (CcO) in the respiratory chain catalyzes oxygen reduction by coupling electron and proton transfer through the enzyme and proton pumping across the membrane. Although the functional unit of CcO is monomeric, mitochondrial CcO forms a monomer and a dimer, as well as a supercomplex with respiratory complexes I and III. A recent study showed that dimeric CcO has lower activity than monomeric CcO and proposed that dimeric CcO is a standby form for enzymatic activation in the mitochondrial membrane. Other studies have suggested that the dimerization is dependent on specifically arranged lipid molecules, peptide segments, and post-translationally modified amino acid residues. To re-examine the structural basis of dimerization, we improved the resolution of the crystallographic structure to 1.3 Å by optimizing the method for cryoprotectant soaking. The observed electron density map revealed many weakly bound detergent and lipid molecules at the interface of the dimer. The dimer interface also contained hydrogen bonds with tightly bound cholate molecules, hydrophobic interactions between the transmembrane helices, and a Met-Met interaction between the extramembrane regions. These results imply that binding of physiological ligands structurally similar to cholate could trigger dimerization in the mitochondrial membrane and that non-specifically bound lipid molecules at the transmembrane surface between monomers support the stabilization of the dimer. The weak interactions involving the transmembrane helices and extramembrane regions may play a role in positioning each monomer at the correct orientation in the dimer.
呼吸链中的细胞色素氧化酶（CcO）通过耦合电子和质子通过该酶的转移以及质子跨膜泵送，催化氧的还原。尽管CcO的功能单元是单体，但线粒体CcO会形成单体、二聚体，以及与呼吸复合物I和III的超复合物。最近的一项研究表明，二聚体CcO的活性低于单体CcO，并提出二聚体CcO是线粒体膜中酶激活的备用形式。其他研究表明，二聚化取决于特定排列的脂质分子、肽段和翻译后修饰的氨基酸残基。为了重新研究二聚化的结构基础，我们通过优化冷冻保护剂浸泡方法，将晶体结构的分辨率提高到了1.3 Å。观察到的电子密度图显示，在二聚体界面处有许多弱结合的去污剂和脂质分子。二聚体界面还包含与紧密结合的胆酸盐分子的氢键、跨膜螺旋之间的疏水相互作用，以及膜外区域之间的甲硫氨酸-甲硫氨酸相互作用。这些结果表明，与胆酸盐结构相似的生理配体的结合可能会触发线粒体膜中的二聚化，并且单体之间跨膜表面的非特异性结合脂质分子支持二聚体的稳定。涉及跨膜螺旋和膜外区域的弱相互作用可能在将每个单体定位在二聚体中的正确方向上起作用。
