A Comprehensive Evaluation of Protein Structure Prediction Models for Short Peptides

Abstract

Short peptides pose distinct challenges for computational structural biology due to their lack of stable tertiary structures, high conformational flexibility, and limited evolutionary signals. To address how modern deep-learning architectures navigate these challenges, we conducted a comprehensive benchmarking of five state-of-the-art protein structure prediction models: AlphaFold2, RoseTTAFold2, ESMFold, OmegaFold, and DMPfold2. Using a curated dataset of experimentally determined short peptide structures (10-49 amino acids) from the Protein Data Bank, we systematically evaluated predictive performance across varying sequence lengths and secondary structure classes. Our results demonstrate that prediction accuracy systematically improves with peptide length. Furthermore, all models perform significantly better on -helical and mixed-structure peptides compared to {beta}-sheet-rich and intrinsically disordered sequences. Among the evaluated methods, AlphaFold2 and the single-sequence language models, ESMFold and Omegafold proved to be the most consistent and accurate overall. We also observed that internal model confidence scores are imperfectly calibrated for short peptides, necessitating cautious interpretation. Finally, by extending our analysis to the dbAMP3 dataset of uncharacterized antimicrobial peptides, we demonstrate that a multi-model consensus approach provides a rational framework for identifying robust structural hypotheses in the absence of experimental reference structures.