Designing antibodies is complex and resource intensive. While deep learning and generative approaches have shown promise in the design of protein binders, achieving high affinity and stability remains challenging. We introduce EvolveX, a structure-based antibody design pipeline leveraging the empirical force field FoldX to design complementarity-determining regions (CDRs) of single-domain antibodies (VHHs). We demonstrate the ability of EvolveX to redesign a VHH targeting mouse Vsig4 (mVsig4) to address two challenges: enhancing stability and affinity for mVsig4 and redesigning it for high affinity to the human ortholog. Notably, EvolveX improved the binding affinity of VHHs to human Vsig4 by over 1,000-fold. Structural analyses by X-ray crystallography confirmed design accuracy. Next-generation sequencing (NGS) analysis further demonstrated the efficiency of FoldX-based design pipeline. Collectively, our study highlights EvolveX's potential to overcome current limitations in antibody design, offering a powerful tool for the development of therapeutics with enhanced specificity, stability, and efficacy.
In this meet-the-author Q&A, Structure's editor-in-chief, Karin Kühnel, speaks to Cathleen Zeymer from the Technical University of Munich about her research group's recent Structure paper entitled "Modular protein scaffold architecture and AI-guided sequence optimization facilitate de novo metalloenzyme engineering" and her work and career.
DNA mismatch repair is an evolutionarily conserved repair pathway that corrects replication errors, thereby preventing genome instability. Two evolutionarily conserved proteins, MutS and MutL, recognize the mismatch and mark the newly synthesized strand for repair. Previous studies have shown how bacterial MutS homodimers function asymmetrically to recognize mismatches and recruit MutL. However, whether MutL homodimers also function asymmetrically to coordinate binding to MutS and activation of their nuclease activity remains unclear. Here, we characterize the ATPase domain of Bacillus subtilis MutL, a MutL protein with endonuclease activity, and delineate the differences with Escherichia coli MutL, a homolog without endonuclease activity. We find that B. subtilis MutL has low affinity for ATP and samples a repertoire of conformations that resemble those observed in eukaryotic MutL paralogs, indicating a relationship between ATP-induced dimer compaction and nuclease activity.
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