EXTRARNAS: A Framework for Extracting RNA Structures with Multiple Tools

Accurate annotation of RNA base-pairing interactions is essential for structural analysis, benchmarking, and data-driven RNA structure prediction. Several tools can extract RNA interactions from three-dimensional coordinates, but their outputs are heterogeneous and may disagree, particularly for non-canonical base pairs. We present EXTRARNAS, a Java-based framework for automated, reproducible, and user-friendly large-scale extraction of RNA structural annotations with multiple tools. EXTRARNAS processes batches of RNA structures specified by PDB identifier and chain, or provided as local PDB files, executes annotation tools through a Docker-based environment, and parses tool-specific outputs using ANTLR4-based grammars. For each structure–tool pair, the framework generates standard BPSEQ files for canonical cis Watson–Crick interactions and introduces BPSEQE, a standardized text format for representing the extended secondary structure, preserving canonical, non-canonical, and multiple interactions per nucleotide. The current prototype supports RNAView, MC-Annotate, and RNAPolis Annotator. We demonstrate EXTRARNAS on eight RNA structures containing triple-helix motifs, comparing extracted canonical pairs against curated BPSEQ references and evaluating the recovery of manually validated Hoogsteen interactions. The results show consistent differences among tools, especially for non-canonical interactions, highlighting the need for standardized representations such as BPSEQE to support reproducible comparison and future consensus-based annotation.