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Abstract
Protein–protein interactions are well-known to regulate enzyme activity in cell signaling and metabolism. Here, we show that protein–protein interactions regulate the activity of a respiratory-chain enzyme, CymA, by changing the direction or bias of catalysis. CymA, a member of the widespread NapC/NirT superfamily, is a menaquinol-7 (MQ-7) dehydrogenase that donates electrons to several distinct terminal reductases in the versatile respiratory network of Shewanella oneidensis. We report the incorporation of CymA within solid-supported membranes that mimic the inner membrane architecture of S. oneidensis. Quartz-crystal microbalance with dissipation (QCM-D) resolved the formation of a stable complex between CymA and one of its native redox partners, flavocytochrome c3 (Fcc3) fumarate reductase. Cyclic voltammetry revealed that CymA alone could only reduce MQ-7, while the CymA-Fcc3 complex catalyzed the reaction required to support anaerobic respiration, the oxidation of MQ-7. We propose that MQ-7 oxidation in CymA is limited by electron transfer to the hemes and that complex formation with Fcc3 facilitates the electron-transfer rate along the heme redox chain. These results reveal a yet unexplored mechanism by which bacteria can regulate multibranched respiratory networks through protein–protein interactions.
Highlights
Protein−protein complexes are fundamental to life where they are key to processes ranging from central metabolism to cell signaling
Cyclic voltammetry revealed that CymA in isolation catalyzed the reduction of MQ-7 but failed to perform the reaction that underpins anaerobic respiration, the oxidation of MQ-7.11 Here, we report the extension of these studies in which the quinone oxidoreductase activity of CymA is determined in the absence and presence of its ET partner, Fcc[3] fumarate reductase.[12]
solid-supported lipid membranes (SSMs) formation was confirmed by a drop in frequency of ∼26− 27 Hz and a small change in dissipation (Figure 2A; for clarity, the formation of the bilayer itself is not shown and the trace starts after the bilayer is formed)
Summary
Protein−protein complexes are fundamental to life where they are key to processes ranging from central metabolism to cell signaling. Transient protein−protein interactions generally underpin the electron-transfer (ET) pathways of respiration.[1] One of the many well-characterized examples of a transient ET complex is that between cytochrome c and cytochrome c oxidase.[2−5] The interaction between these partner proteins is weak and dynamic. This ensures the frequent exchange of partner proteins as required to support electron flux in cases where the sole function of one of the proteins is to shuttle electrons between redox partners.[1] While it is generally assumed that such transient protein−protein interactions are specific, for Paracoccus denitrificans it has recently been shown that seven proteins in a respiratory network interact in a seemingly ill-defined manner.[6] This results in an intricate electron-transfer network that may be better suited to successful colonization of habitats with changing resources
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