biostudies-literatureUnknown16(1)Huang QChangping LaboratoryStabilizing the RSV F protein in its prefusion conformation is crucial for effective vaccine development but has remained a significant challenge. Traditional stabilization methods, such as disulfide bonds and cavity-filling mutations, have been labor-intensive and have often resulted in suboptimal expression levels. Here, we report the design of an RSV prefusion F (preF) antigen using a proline-scanning strategy, incorporating seven proline substitutions to achieve stabilization. The resulting variant, preF7P, is structurally and biochemically validated to maintain the correct prefusion state. PreF7P demonstrates superior immunogenicity with a 1.8-fold increase in neutralizing antibody titers when compared to DS-cav2, and provides protection from clinical disease against both RSV A and B Nature communications7805https://www.ebi.ac.uk/biostudies/studies/S-EPMC12370985biostudies-literatureHighly scalable prefusion-stabilized RSV F vaccine with enhanced immunogenicity and robust protection.PMC12370985Yan JLang QWang FHan XHuang QBai YGao CLi YZhao LCao XBian LLuo LWang YDuan XSong YXu YfalseHighly scalable prefusion-stabilized RSV F vaccine with enhanced immunogenicity and robust protection.Stabilizing the RSV F protein in its prefusion conformation is crucial for effective vaccine development but has remained a significant challenge. Traditional stabilization methods, such as disulfide bonds and cavity-filling mutations, have been labor-intensive and have often resulted in suboptimal expression levels. Here, we report the design of an RSV prefusion F (preF) antigen using a proline-scanning strategy, incorporating seven proline substitutions to achieve stabilization. The resulting variant, preF7P, is structurally and biochemically validated to maintain the correct prefusion state. PreF7P demonstrates superior immunogenicity with a 1.8-fold increase in neutralizing antibody titers when compared to DS-cav2, and provides protection from clinical disease against both RSV A and B 2025-01-01T00:00:00Z2025 Aug2026-05-08T10:52:10.894Z2026-05-03T03:06:03.485ZS-EPMC123709854084137210.1038/s41467-025-63084-z