Molecular Mechanism of STIL Coiled-Coil Domain Oligomerization

Autor: Mai Shamir, Freddie J. O. Martin, Derek N. Woolfson, Assaf Friedler
Jazyk: angličtina
Rok vydání: 2023
Předmět:
Zdroj: International Journal of Molecular Sciences, Vol 24, Iss 19, p 14616 (2023)
Druh dokumentu: article
ISSN: 1422-0067
1661-6596
DOI: 10.3390/ijms241914616
Popis: Coiled-coil domains (CCDs) play key roles in regulating both healthy cellular processes and the pathogenesis of various diseases by controlling protein self-association and protein–protein interactions. Here, we probe the mechanism of oligomerization of a peptide representing the CCD of the STIL protein, a tetrameric multi-domain protein that is over-expressed in several cancers and associated with metastatic spread. STIL tetramerization is mediated both by an intrinsically disordered domain (STIL400–700) and a structured CCD (STIL CCD718–749). Disrupting STIL oligomerization via the CCD inhibits its activity in vivo. We describe a comprehensive biophysical and structural characterization of the concentration-dependent oligomerization of STIL CCD peptide. We combine analytical ultracentrifugation, fluorescence and circular dichroism spectroscopy to probe the STIL CCD peptide assembly in solution and determine dissociation constants of both the dimerization, (KD = 8 ± 2 µM) and tetramerization (KD = 68 ± 2 µM) of the WT STIL CCD peptide. The higher-order oligomers result in increased thermal stability and cooperativity of association. We suggest that this complex oligomerization mechanism regulates the activated levels of STIL in the cell and during centriole duplication. In addition, we present X-ray crystal structures for the CCD containing destabilising (L736E) and stabilising (Q729L) mutations, which reveal dimeric and tetrameric antiparallel coiled-coil structures, respectively. Overall, this study offers a basis for understanding the structural molecular biology of the STIL protein, and how it might be targeted to discover anti-cancer reagents.
Databáze: Directory of Open Access Journals
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