ENSURE: the encyclopedia of suppressor tRNA with an AI assistant

Suppressor transfer RNAs (sup-tRNAs) offer a promising strategy for rescuing proteins truncated by premature termination codons via translational readthrough. Despite recent advances in genetic code expansion and RNA therapeutics that have facilitated sup-tRNA engineering, progress remains constrained by the lack of a dedicated, integrative data platform. Here, we present ENSURE (https://trna.lumoxuan.cn/), a comprehensive knowledge base that aggregates 2152 disease- and cancer-associated nonsense, missense, and frameshift variants; 86 experimentally validated natural sup-tRNAs, 1108 tRNA engineering strategies; and 487 curated tRNA element records. All sequences undergo multiple sequence alignment, secondary structure prediction, and AlphaFold 3 modeling accompanied by interactive 2D/3D visualization. ENSURE supports keyword and BLAST searches, as well as bulk downloads. A key feature is an AI assistant based on a retrieval-augmented generation architecture: ~123.7k tokens of database pages, literature abstracts, structural annotations, and other resources are chunked and encoded with Sentence-BERT; user queries are processed similarly, matched to relevant chunks, concatenated with the query, and passed to a large language model to generate answers with illustrative resources inserted automatically. By combining curated data, structural models, and an interactive AI assistant, ENSURE provides a powerful platform to accelerate sup-tRNA research and translational applications.