Browsing by Author "Gonzalez-Higueras, Jorge"

Now showing 1 - 3 of 3
  • No Thumbnail Available
    Item
    A contact-based analysis of local energetic frustration dynamics identifies key residues enabling RfaH fold-switch
    (2024) Gonzalez-Higueras, Jorge; Freiberger, Maria Ines; Galaz-Davison, Pablo; Parra, R. Gonzalo; Ramirez-Sarmiento, Cesar A.
    Fold-switching enables metamorphic proteins to reversibly interconvert between two highly dissimilar native states to regulate their protein functions. While about 100 proteins have been identified to undergo fold-switching, unveiling the key residues behind this mechanism for each protein remains challenging. Reasoning that fold-switching in proteins is driven by dynamic changes in local energetic frustration, we combined fold-switching simulations generated using simplified structure-based models with frustration analysis to identify key residues involved in this process based on the change in the density of minimally frustrated contacts during refolding. Using this approach to analyze the fold-switch of the bacterial transcription factor RfaH, we identified 20 residues that significantly change their frustration during its fold-switch, some of which have been experimentally and computationally reported in previous works. Our approach, which we developed as an additional module for the FrustratometeR package, highlights the role of local frustration dynamics in protein fold-switching and offers a robust tool to enhance our understanding of other proteins with significant conformational shifts.
  • No Thumbnail Available
    Item
    Concerted transformation of a hyper-paused transcription complex and its reinforcing protein
    (2024) Zuber, Philipp K.; Said, Nelly; Hilal, Tarek; Wang, Bing; Loll, Bernhard; Gonzalez-Higueras, Jorge; Ramirez-Sarmiento, Cesar A.; Belogurov, Georgiy A.; Artsimovitch, Irina; Wahl, Markus C.; Knauer, Stefan H.
    RfaH, a paralog of the universally conserved NusG, binds to RNA polymerases (RNAP) and ribosomes to activate expression of virulence genes. In free, autoinhibited RfaH, an alpha-helical KOW domain sequesters the RNAP-binding site. Upon recruitment to RNAP paused at an ops site, KOW is released and refolds into a beta-barrel, which binds the ribosome. Here, we report structures of ops-paused transcription elongation complexes alone and bound to the autoinhibited and activated RfaH, which reveal swiveled, pre-translocated pause states stabilized by an ops hairpin in the non-template DNA. Autoinhibited RfaH binds and twists the ops hairpin, expanding the RNA:DNA hybrid to 11 base pairs and triggering the KOW release. Once activated, RfaH hyper-stabilizes the pause, which thus requires anti-backtracking factors for escape. Our results suggest that the entire RfaH cycle is solely determined by the ops and RfaH sequences and provide insights into mechanisms of recruitment and metamorphosis of NusG homologs across all life.
  • No Thumbnail Available
    Item
    DNA facilitates heterodimerization between human transcription factors FoxP1 and FoxP2 by increasing their conformational flexibility
    (2023) Conuecar, Ricardo; Asela, Isabel; Rivera, Maira; Galaz-Davison, Pablo; Gonzalez-Higueras, Jorge; Hamilton, George L.; Engelberger, Felipe; Ramirez-Sarmiento, Cesar A.; Babul, Jorge; Sanabria, Hugo; Medina, Exequiel
    Transcription factors regulate gene expression by binding to DNA. They have disordered regions and specific DNA-binding domains. Binding to DNA causes structural changes, including folding and interactions with other molecules. The FoxP subfamily of transcription factors in humans is unique because they can form heterotypic interactions without DNA. However, it is unclear how they form heterodimers and how DNA binding affects their function. We used computational and experimental methods to study the structural changes in FoxP1's DNA-binding domain when it forms a heterodimer with FoxP2. We found that FoxP1 has complex and diverse conformational dynamics, transitioning between compact and extended states. Surprisingly, DNA binding increases the flexibility of FoxP1, contrary to the typical folding-upon-binding mechanism. In addition, we observed a 3-fold increase in the rate of heterodimerization after FoxP1 binds to DNA. These findings emphasize the importance of structural flexibility in promoting heterodimerization to form transcriptional complexes.