Abstract
Flavin is covalently attached to the protein scaffold in ~10% of flavoenzymes. However, the mechanism of covalent modification is unclear, due in part to challenges in stabilizing assembly intermediates. Here, we capture the structure of an assembly intermediate of the Escherichiacoli Complex II (quinol:fumarate reductase (FrdABCD)). The structure contains the E. coli FrdA subunit bound to covalent FAD and crosslinked with its assembly factor, SdhE. The structure contains two global conformational changes as compared to prior structures of the mature protein: the rotation of a domain within the FrdA subunit, and the destabilization of two large loops of the FrdA subunit, which may create a tunnel to the active site. We infer a mechanism for covalent flavinylation. As supported by spectroscopic and kinetic analyses, we suggest that SdhE shifts the conformational equilibrium of the FrdA active site to disfavor succinate/fumarate interconversion and enhance covalent flavinylation.
Highlights
Flavin is covalently attached to the protein scaffold in ~10% of flavoenzymes
We developed methods to crosslink the complex between the FrdA subunit of E. coli Complex II FrdABCD and its assembly factor SdhE19
In evaluating the global architecture of this FrdA-SdhE assembly intermediate, the SdhE binding site is located on a surface of the FrdA subunit that interacts with the FrdB subunit in the assembled FrdABCD complex[17] (Fig. 1a–c)
Summary
Flavin is covalently attached to the protein scaffold in ~10% of flavoenzymes. the mechanism of covalent modification is unclear, due in part to challenges in stabilizing assembly intermediates. Integral-membrane Complex II enzymes can act in either aerobic respiration (termed succinate:quinone oxidoreductase (SdhABCD)) or during anaerobic respiration with fumarate as the terminal electron acceptor (termed quinol: fumarate reductase (FrdABCD)) In their mature and fully assembled forms, both SdhABCD and FrdABCD can catalyze the interconversion of fumarate and succinate at flavin adenine dinucleotide (FAD) covalently attached to the flavoprotein subunit (FrdA or SdhA). One assembly factor (termed SdhAF2 in humans, SdhE in Escherichia coli, and Sdh[5] in yeast) enhances covalent flavinylation in both human and bacterial Complex II homologs[5] This small protein (~90–140 amino acids, depending on the organism) is conserved in all kingdoms[5,8,9]
Sign-in/Register to view highlights & summary Sign-in/Register to access full text options 
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a callDisclaimer: All third-party content on this website/platform is and will remain the property of their respective owners and is provided on "as is" basis without any warranties, express or implied. Use of third-party content does not indicate any affiliation, sponsorship with or endorsement by them. Any references to third-party content is to identify the corresponding services and shall be considered fair use under The CopyrightLaw.
