Multi-site-specific isotopic labeling accelerates high-resolution structural investigations of pathogenic huntingtin exon-1

• Published in: Structure (London) Vol. 31; no. 6; pp. 644 - 650.e5
• Main Authors: Elena-Real, Carlos A., Urbanek, Annika, Lund, Xamuel L., Morató, Anna, Sagar, Amin, Fournet, Aurélie, Estaña, Alejandro, Bellande, Tracy, Allemand, Frédéric, Cortés, Juan, Sibille, Nathalie, Melki, Ronald, Bernadó, Pau
• Format: Journal Article
• Language: English
• Published: United States Elsevier Ltd 01-06-2023; Elsevier (Cell Press)
• Subjects: Biochemistry, Molecular Biology; Bioinformatics; Computer Science; conformational ensemble; huntingtin; Huntington’s disease; intrinsically disordered proteins; isotopic labeling; Life Sciences; nuclear magnetic resonance; poly-Q; Structural Biology; tRNA suppression; isotopic labeling; conformational ensemble; huntingtin; Huntington’s disease; tRNA suppression; nuclear magnetic resonance; poly-Q; intrinsically disordered proteins
• DOI: 10.1016/j.str.2023.04.003
• EISSN: 1878-4186
• ISSN: 0969-2126

Huntington’s disease neurodegeneration occurs when the number of consecutive glutamines in the huntingtin exon-1 (HTTExon1) exceeds a pathological threshold of 35. The sequence homogeneity of HTTExon1 reduces the signal dispersion in NMR spectra, hampering its structural characterization. By simultaneously introducing three isotopically labeled glutamines in a site-specific manner in multiple concatenated samples, 18 glutamines of a pathogenic HTTExon1 with 36 glutamines were unambiguously assigned. Chemical shift analyses indicate the α-helical persistence in the homorepeat and the absence of an emerging toxic conformation around the pathological threshold. Using the same type of samples, the recognition mechanism of Hsc70 molecular chaperone has been investigated, indicating that it binds to the N17 region of HTTExon1, inducing the partial unfolding of the poly-Q. The proposed strategy facilitates high-resolution structural and functional studies in low-complexity regions.
[Display omitted]
•Three isotopically labeled glutamines were site-specifically introduced in huntingtin•A huntingtin construct with a 36-residue-long poly-Q was assigned using this approach•Absence of a toxic conformation at the pathological threshold of Huntington’s disease•Chaperone Hsc70 interacts with N17 and unfolds the α-helical structure of the poly-Q
Using nine concatenated samples with three isotopically labeled glutamines, Elena-Real et al. have structurally characterized the poly-Q region of a pathogenic huntingtin exon-1 containing 36 glutamines, indicating that no toxic conformation emerges at the Huntington’s disease pathological threshold. The structural impact on huntingtin upon binding of the chaperone Hsc70 was unveiled.