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Sperl, L.E.* ; Hagn, F.

NMR structural and biophysical analysis of the disease-linked inner mitochondrial membrane protein MPV17.

J. Mol. Biol. 433:167098 (2021)
DOI
Open Access Green: Postprint online verfügbar 07/2022
MPV17 is an integral inner mitochondrial membrane protein, whose loss-of-function is linked to the hepatocerebral form of the mitochondrial-DNA-depletion syndrome, leading to a tissue-specific reduction of mitochondrial DNA and organ failure in infants. Several disease-causing mutations in MPV17 have been identified and earlier studies with reconstituted protein suggest that MPV17 forms a high conductivity channel in the membrane. However, the molecular and structural basis of the MPV17 functionality remain only poorly understood. In order to make MPV17 accessible to high-resolution structural studies, we here present an efficient protocol for its high-level production in E. coli and refolding into detergent micelles. Using biophysical and NMR methods, we show that refolded MPV17 in detergent micelles adopts a compact structure consisting of six membrane-embedded α-helices. Furthermore, we demonstrate that MPV17 forms oligomers in a lipid bilayer that are further stabilized by disulfide-bridges. In line with these findings, MPV17 could only be inserted into lipid nanodiscs of 8-12 nm in diameter if intrinsic cysteines were either removed by mutagenesis or blocked by chemical modification. Using this nanodisc reconstitution approach, we could show that disease-linked mutations in MPV17 abolish its oligomerization properties in the membrane. These data suggest that, induced by oxidative stress, MPV17 can alter its oligomeric state from a properly folded monomer to a disulfide-stabilized oligomeric pore which might be required for the transport of metabolic DNA precursors into the mitochondrial matrix to compensate for the damage caused by reactive oxygen species.
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Publikationstyp Artikel: Journalartikel
Dokumenttyp Wissenschaftlicher Artikel
Schlagwörter Dynamics ; Membrane Protein Stability ; Mitochondrial Diseases ; Nanodiscs ; Reactive Oxygen Species; Phospholipid-bilayer Nanodiscs; Kidney-disease; Gene Mpv17; Encodes; Channel; Sym1; Reconstruction; Resolution; Ortholog
ISSN (print) / ISBN 0022-2836
e-ISSN 1089-8638
Quellenangaben Band: 433, Heft: 15, Seiten: , Artikelnummer: 167098 Supplement: ,
Verlag Elsevier
Verlagsort 24-28 Oval Rd, London Nw1 7dx, England
Begutachtungsstatus Peer reviewed
Förderungen Center for Integrated Protein Science Munich (CIPSM)
European Union
Technical University of Munich, Institute for Advanced Study - German Excellence Initiative
Helmholtz Society