Disulfide-constrained Fabs overcome target size limitation for high-resolution single particle cryoEM.

ABSTRACT: High-resolution protein structures are essential for understanding biological mechanisms and drug discovery. While cryoEM has revolutionized structure determination of large protein complexes, most disease-related proteins are small (<50 kDa) and challenging to resolve due to low signal-to-noise ratios and alignment difficulties. Current scaffold protein strategies increase target size but suffer from inherent flexibility, resulting in poorly resolved targets compared to scaffolds. We present an iteratively engineered molecular design transforming antibody fragments (Fabs) into conformationally Rigid Fabs that enable high-resolution structure determination of small proteins (~20 kDa). This design introduces strategic disulfide bonds, creating well-folded, rigidly constrained Fabs applicabl

SUBMITTER: Kung JE 

PROVIDER: S-EPMC12484979 | biostudies-literature | 2025 Sep

REPOSITORIES: biostudies-literature