Structural and Computational Analysis of the Quinone-binding Site of Complex II (Succinate-Ubiquinone Oxidoreductase)

Abstract

The transfer of electrons and protons between membranebound respiratory complexes is facilitated by lipid-soluble redox-active quinone molecules (Q). This work presents a structural analysis of the quinone-binding site (Q-site) identified in succinate:ubiquinone oxidoreductase (SQR) from Escherichia coli. SQR, often referred to as Complex II or succinate dehydrogenase, is a functional member of the Krebs cycle and the aerobic respiratory chain and couples the oxidation of succinate to fumarate with the reduction of quinone to quinol (QH 2 ). The interaction between ubiquinone and the Q-site of the protein appears to be mediated solely by hydrogen bonding between the O1 carbonyl group of the quinone and the side chain of a conserved tyrosine residue. In this work, SQR was co-crystallized with the ubiquinone binding-site inhibitor Atpenin A5 (AA5) to confirm the binding position of the inhibitor and reveal additional structural details of the Q-site. The electron density for AA5 was located within the same hydrophobic pocket as ubiquinone at, however, a different position within the pocket. AA5 was bound deeper into the site prompting further assessment using proteinligand docking experiments in silico. The initial interpretation of the Q-site was re-evaluated in the light of the new SQR-AA5 structure and protein-ligand docking data. Two binding positions, the Q 1 -site and Q 2 -site, are proposed for the E. coli SQR quinone-binding site to explain these data. At the Q 2 -site, the side chains of a serine and histidine residue are suitably positioned to provide hydrogen bonding partners to the O4 carbonyl and methoxy groups of ubiquinone, respectively. This allows us to propose a mechanism for the reduction of ubiquinone during the catalytic turnover of the enzyme.The Complex II family is comprised of two homologous integral membrane proteins (1-4). Succinate:quinone oxidoreductase (SQR), 4 or succinate dehydrogenase (SDH), is a functional member of the Krebs cycle and the aerobic respiratory chain coupling the oxidation of succinate to fumarate with the reduction of quinone (Q) to quinol (QH 2 ). Quinol:fumarate oxidoreductase (QFR), or fumarate reductase, catalyzes the reverse reaction to SQR during anaerobic respiration. Both enzymes have a similar subunit and cofactor composition (1-4). The hydrophilic subunits, composed of a flavoprotein (SdhA) and an ironsulfur protein (SdhB), have a high degree of sequence homology across species. The SdhAB catalytic subunits are anchored to the membrane by the hydrophobic SdhCD subunits, which in the case of mammalian and Escherichia coli Complex II form a cytochrome b that contains one heme b. The two-electron oxidation of succinate at the substrate binding site in the SdhA subunit is coupled to the two-electron reduction of quinone in the membrane domain of Complex II. Electrons are sequentially transferred to ubiquinone (in the case of mammalian and E. coli Quinones can undergo 2e Ϫ /2H ϩ oxidation-reduction reactions, an essential feature of respiration, allowing the t...

Plasticity of the Quinone-binding Site of the Complex II Homolog Quinol:Fumarate Reductase

“…The results shown in Tables 2 and 3 parallel previous findings with E. coli SQR where site-directed substitutions near the UQbinding site did not identify any single amino acid side chain that was essential for proton shuttling to UQ (37)(38)(39). Studies on SQR have included the mutation of side chains that stabilize ordered water molecules leading into the UQ-binding site, and the substitution of these has measurable but limited effects on quinone reduction (38).…”

Enantioselective total synthesis of atpenin A5

“…Furthermore, the crystal structure analysis of Escherichia coli complex II-atpenin A5 (3) complex has been achieved, and catalytic reduction of quinone was suggested to occur at the atpenin-binding site of E. coli complex II. 6 As described, atpenins are expected to be useful as biochemical tools in the molecular-biological research of complex II. We report herein the enantioselective total synthesis of atpenin A5 (3) by a convergent strategy through a coupling reaction between 2,3,4,6-tetraalkoxypyridine (4a) and a side-chain aldehyde (5), as shown in Figure 2.…”

Why the Flavin Adenine Dinucleotide (FAD) Cofactor Needs To Be Covalently Linked to Complex II of the Electron‐Transport Chain for the Conversion of FADH2 into FAD

“…This water channel crosses the membrane anchor and arrives at the ubiquinone binding site 8a. It is probable that the eukaryotic complex has also a water channel for proton uptake.…”