The giant sarcomere protein titin is the third filament in muscle and is integral to maintaining sarcomere integrity as well as contributing to both active and passive tension. Titin is a multi-domain protein that contains regions of repeated structural elements. The N2A region sits at the boundary between the proximal Ig region of titin that is extended under low force and the PEVK region that is extended under high force. Multiple binding interactions have been associated with the N2A region and it has been proposed that this region acts as a mechanical stretch sensor. The focus of this work is a 117 amino acid portion of the N2A region (N2A-IS), which resides between the proximal Ig domains and the PEVK region. Our work has shown that the N2A-IS region is predicted to contain helical structure in the center while both termini are predicted to be disordered. Recombinantly expressed N2A-IS protein contains 13% α-helical structure, as measured via circular dichroism. Additional α-helical structure can be induced with 2,2,2-trifluoroethanol, suggesting that there is transient helical structure that might be stabilized in the context of the entire N2A region. The N2A-IS region does not exhibit any cooperativity in either thermal or chemical denaturation studies while size exclusion chromatography and Fluorescence Resonance Energy Transfer demonstrates that the N2A-IS region has an extended structure. Combined, these results lead to a model of the N2A-IS region having a helical core with extended N- and C-termini.
|Original language||English (US)|
|Number of pages||10|
|Journal||Biochimica et Biophysica Acta - Proteins and Proteomics|
|State||Published - Jan 1 2017|
- Disordered region
- Extended structure
- Secondary structure
ASJC Scopus subject areas
- Analytical Chemistry
- Molecular Biology
The insertion sequence of the N2A region of titin exists in an extended structure with helical characteristics. / Tiffany, Holly; Sonkar, Kanchan; Gage, Matthew J.In: Biochimica et Biophysica Acta - Proteins and Proteomics, Vol. 1865, No. 1, 01.01.2017, p. 1-10.
Research output: Contribution to journal › Article