Structural basis for the regulatory mechanism of mammalian mitochondrial respiratory chain megacomplex-Ⅰ_(2)Ⅲ_(2)Ⅳ_(2)

Mammalian mitochondrial electron transport chain complexes are the most important and complicated protein machinery in mitochondria.Although this system has been studied for more than a century,its composition and molecular mechanism are still largely unknown.Here we report the high-resolution cryo-electron microscopy(Cryo-EM)structures of porcine respiratory chain megacomplex-Ⅰ_(2)Ⅲ_(2)Ⅳ_(2)(MCⅠ_(2)Ⅲ_(2)Ⅳ_(2))in five different conformations,including State 1,State 2,Mid 1,Mid 2,and Mid 3.High-resolution Cryo-EM imaging,combined with super-resolution gated stimulated emission depletion microscopy(gSTED),strongly supports the formation of MCⅠ_(2)Ⅲ_(2)Ⅳ_(2)in live cells.Each MCⅠ_(2)Ⅲ_(2)Ⅳ_(2)structure contains 141 subunits(70 different kinds of peptides,2.9 MDa)in total with 240 transmembrane helices.The mutual influence among CⅠ,CⅢ,and CⅣshown in the MCⅠ_(2)Ⅲ_(2)Ⅳ_(2)structure suggests this megacomplex could act as an integral unit in electron transfer and proton pumping.The conformational changes from different states suggest a plausible regulatory mechanism for the MCⅠ_(2)Ⅲ_(2)Ⅳ_(2)activation/deactivation process.