Cryo-EM structures of the air-oxidized and dithionite-reduced photosynthetic alternative complex III from Roseiflexus castenholzii

Abstract

Alternative complex III (ACIII) is a multisubunit quinol:electron acceptor oxidoreductase that couples quinol oxidation with transmembrane proton translocation in both the respiratory and photosynthetic electron transport chains of bacteria. The coupling mechanism, however, is poorly understood. Here, we report the cryo-EM structures of air-oxidized and dithionite-reduced ACIII from the photosynthetic bacterium Roseiflexus castenholzii at 3.3- and 3.5-Å resolution, respectively. We identified a menaquinol binding pocket and an electron transfer wire comprising six hemes and four iron-sulfur clusters that is capable of transferring electrons to periplasmic acceptors. We detected a proton translocation passage in which three strictly conserved, mid-passage residues are likely essential for coupling the redox-driven proton translocation across the membrane. These results allow us to propose a previously unrecognized coupling mechanism that links the respiratory and photosynthetic functions of ACIII. This study provides a structural basis for further investigation of the energy transformation mechanisms in bacterial photosynthesis and respiration.

Ionogel‐Electrode for the Study of Protein Tunnel Junctions under Physiologically Relevant Conditions

“…The phenomenon of tunneling across biomolecules is associated with numerous fundamental studies and electronic applications, for examples, the pathways of electrons in biosystem [1][2][3] , electronic biomaterials 4,5 , and bio(semi-)conductors [6][7][8] . In particular, the investigation of tunneling through proteins is important for us to understand the protein associated electrochemical processes, for instance respiration 9,10 , photosynthesis 11,12 and biocatalytic reactions 13,14 . Two types of tunneling are found in protein junctions: The coherent tunneling process is temperature inactive electron transport (ETp) meaning that the protein with its complex structure only works as a certain tunneling barrier between electrodes; The incoherent tunneling is temperature active electron transfer (ET) often involving hopping of electron between electrodes and redox-active center with a certain activation energy (E a ).…”

The Monoheme c Subunit of Respiratory Alternative Complex III Is Not Essential for Electron Transfer to Cytochrome aa 3 in Flavobacterium johnsoniae

“…Recently, ACIII structures from three species, Flavobacterium johnsoniae (PDB: 6BTM ), R. marinus (PDB: 6F0K ), and Roseiflexus castenholzii (PDB: 6LOE and 6LOD ) were solved by cryo-EM, revealing no structural similarity to cytochrome bc 1 / b 6 f ( 6 – 8 ). Based on the sequence similarities, ACIII is considered to be a new member of the complex iron-sulfur molybdoenzyme (CISM) superfamily, but without the molybdenum cofactor ( 2 , 5 ).…”

“…ActE is a membrane-anchored subunit that contains one heme c ( 6 ). R. marinus and R. castenholzii ACIII structures are highly similar, with the exception that the ActA subunit lacks one of the hemes ( 7 , 8 ) that is present in F. johnsoniae . Interestingly, unlike in R. castenholzii , ACIII from F. johnsoniae and R. marinus was isolated in a complex with an aa 3 -type oxygen reductase (cytochrome aa 3 in F. johnsoniae and cytochrome caa 3 in R. marinus ) and it was in these forms that they were analyzed by cryo-EM.…”

“…On the contrary, for R. castenholzii , it was postulated that the [4Fe-4S] clusters form a dead end. These clusters may serve as an electron sink limiting formation of reactive oxygen species ( 8 ). However, the actual roles of these clusters remain unknown.…”

“…R. castenholzii ACIII is engaged in cyclic electron transfer during photosynthesis. Electrons from ActE are transferred to the small blue copper protein auracyanin, and then to the photosynthetic reaction center ( 8 ).…”