Oligomerization, trans-reduction, and instability of mutant NOTCH3 in inherited vascular dementia
Year: 2022
Type: article
Abstract: Cerebral small vessel disease (SVD) is a prevalent disease of aging and a major contributor to stroke and dementia. The most commonly inherited SVD, CADASIL, is caused by dominantly acting cysteine-altering mutations in NOTCH3. These mutations change the number of cysteines from an even to an odd number, but the impact of these alterations on NOTCH3 protein structure remain unclear. Here, we prepared wildtype and four mutant recombinant NOTCH3 protein fragments to analyze the impact of CADASIL mutations on oligomerization, thiol status, and protein stability. Using gel electrophoresis, tandem MS/MS, and collision-induced unfolding, we find that NOTCH3 mutant proteins feature increased amounts of inappropriate disulfide bridges, reduced cysteines, and structural instability. Presence of a second protein factor, an N-terminal fragment of NOTCH3 (NTF), is capable of further altering disulfide statuses of both wildtype and mutant proteins, leading to increased numbers of reduced cysteines and further destabilization of NOTCH3 structure. In sum, these studies identify specific cysteine residues alterations and quaternary structure induced by CADASIL mutations in NOTCH3; further, we validate that reductive factors alter the structure and stability of this small vessel disease protein.
Source: Communications Biology
Language: English
Subfield: Neurology
Field: Medicine
Domain: Health Sciences
Open Access status: gold
Estimated APC paid: $3,490
Funders: Foundation for the National Institutes of Health, U.S. Department of Veterans Affairs, American Heart Association, University of Michigan, National Institutes of Health, National Heart, Lung, and Blood Institute, National Cancer Institute, National Institute of General Medical Sciences, National Institute of Neurological Disorders and Stroke
Awards: G1330204347, G1612408122, G1618614147, Integrating Quality Control: Studies of UBQLN2 in Age-Related Neurodegeneration, G2953809093, G3010933817, G3283720050, G3549828030, Training Program in Translational Cardiovascular Research and Entrepreneurship, G4007149577, G4232504776, G4376152923, G4513335875, G4667052213, NOTCH3 N-terminal fragmentation in cerebral small vessel disease, Promoting Diversity of Future Scientists, G5949955087, G6674348244, G6725671518, Identifying the Role of a Pathological Protein Conformation in CADASIL, G8442533357, G8467019912, G8901171743, G9000031400, G9000088481, G9000133651, G9000187910, G9000191659, G9000220969, G9000347736
OpenAlex ID: W4223609027