CryoSHIELDs: tunable amphipathic peptide additives for cryo-EM

Abstract

While cryogenic electron microscopy (cryo-EM) has transformed structural biology, sample preparation remains a major determinant of success. Specifically, the air-water interface (AWI) can damage complexes, impose preferred orientation, and reduce particle density. Here, we introduce cryoSHIELDs, a library of peptide additives designed to modulate the AWI and improve cryo-EM grid preparation. Across 24 diverse protein systems, cryoSHIELDs improved particle distribution, angular sampling, complex integrity, and, in selected cases, map quality and resolution. CryoSHIELDs enabled structure determination of the OSMR{beta}-Vixarelimab complex, the multi-subunit CCAN kinetochore complex without chemical cross-linking, and membrane proteins including ABCC6 and the -opioid receptor-Gi complex, the latter resolved at 2.3 A using tenfold lower protein concentration without compromising map quality. CryoSHIELDs provide a simple, tunable, and broadly applicable platform for more reliable cryo-EM in discovery biology and structure-based drug development.