A high-throughput selection system for fast-acting covalent protein drugs.

Fan, Qiongxuan; Mei, Jiahao; Li, Tian; Zang, Chuanlong; Li, Mengjiao; Tang, Jing; Xu, You; Yu, Ge et al. · Science · 2026

basic_science · Level V

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Abstract

Covalent protein drugs offer therapeutic potential but are limited by slow target engagement and the absence of high-throughput selection platforms. Rapid covalent binding requires coordinated optimization of affinity, stability, and warhead geometry-an intrinsically multidimensional challenge. We develop a yeast display platform coupled with chemoselective modification that enables selection of fast-acting covalent proteins without increasing intrinsic warhead reactivity. Using this system, we engineered a covalent programmed death-ligand 1 (PD-L1) antagonistic nanobody with rapid crosslinking kinetics (<i>k</i><sub>obs</sub> = 0.18 min<sup>-1</sup>, t<sub>1/2</sub> = 3.8 min) and improved tumor suppression compared with envafolimab and atezolizumab. Similarly, we engineered a fast-acting covalent interleukin-18 (IL-18) (<i>k</i><sub>obs</sub> = 0.54 min<sup>-1</sup>, t<sub>1/2</sub> = 1.3 min) and a covalent miniprotein targeting the receptor binding domain (RBD) of SARS-CoV-2, demonstrating applicability across protein modalities.