همایش ملی بیوانفورماتیک ایران

صفحه اصلی / 4th international edition and 13th Iranian Conference on Bioinformatics

Tailoring Stability of Keratinocyte Growth Factor Variants for Effective Wound Healing in Acidic Conditions

نویسندگان :

Mansoureh Shahbazi Dastjerdeh¹ Mohammadtaghi Borjian Boroujeni²

1- Blood Transfusion Research Center, High Institute for Research and Education in Transfusion Medicine, Tehran, Iran 2- Department of Molecular Medicine, Biotechnology Research Center, Pasteur Institute of Iran, Tehran, Iran

کلمات کلیدی :

Fibroblast Growth Factor 7،Wound healing،Protein Stability،Protein Engineering،Molecular Dynamics Simulation

چکیده :

With the global increasing incidence of wound, the prevalence of chronic, difficult-to-heal or non-healing wounds is expected to increase (Holl, Kowalewski et al. 2021). The global wound care market has been valued at $23.66 billion in 2023, $24.72 billion in 2024, and is predicted to reach to USD $38.39 billion in 2034 with a CAGR of 4.5% (Research 2024). The growing understanding of wound healing mechanisms has shed light on the key regulators of this process including surface skin pH (Holl, Kowalewski et al. 2021). pH is one of the critical factors affecting all cellular and biochemical reactions in the wound healing process including angiogenesis, oxygen release, protease activity, and bacterial toxicity (Gethin 2007). It has been demonstrated that the pH of normal skin is in the range of 4.0-6.0 as a critical aspect of its barrier function to prevent infection. However, the wound disrupts the skin pH making an alkaline environment substantially slowing the healing process in chronic wounds (Jones, Cochrane et al. 2015). Keratinocyte Growth Factor (KGF) plays a crucial role in promoting epithelial cell proliferation, making it a valuable candidate for wound healing therapies (Bártolo, Reis et al. 2022). KGF application in the early stages of the wound, accelerates the healing process and prevents the wound from becoming chronic. However, its instability in acidic environments limits its effectiveness in early-stage wound care (Chen, Arakawa et al. 1994, Treuheit, Dharmavaram et al. 2012). The present study addresses the instability of recombinant human Keratinocyte Growth Factor (rhKGF) in acidic environments. Using pH molecular dynamics (pH-MD) simulations, we previously investigated the influence of pH on rhKGF stability and identified positively charged residues as contributors to its instability (Boroujeni, Dastjerdeh et al. 2021). Building on these findings, we employed a rational design strategy to lower KGF's isoelectric point (pI) by introducing targeted mutations. This led to the development of 15 mutants which of them two mutants, K76E and K126E, were selected based on molecular dynamics simulations, and docking studies. Further experimental evaluations including secondary structure analysis, thermal stability, acidic pH stability, and mitogenic activity were performed on these two mutants. According to the results, both mutants retained similar structural characteristics and potency and showed lower thermal stability, but modest acidic pH stability improvements compared to native rhKGF. K126E exhibited enhanced stability over extended periods in acidic environments, indicating its potential as a viable alternative for topical applications. Our findings highlight the efficacy of targeted mutations in acidic pH stability enhancement and underscore the role of electrostatic interactions in KGF stability. Future research should focus on targeting other hotspots for mutants, further computational analysis, and optimizing formulations that leverage these mutants for enhanced therapeutic efficacy in wound care applications.