Fold first, ask later: structure-informed function annotation of Pseudomonas phage proteins

bioRxiv (Cold Spring Harbor Laboratory)

Abstract

Abstract Phages, the viruses of bacteria, harbor an incredibly diverse repertoire of proteins capable of manipulating their bacterial hosts, inspiring many medical and biotechnological applications. However, to date, only a limited subset of that repertoire can be exploited, due to the difficulties in functionally elucidating these proteins. In this study, we investigated several structure-informed approaches to annotate hypothetical proteins from Pseudomonas infecting phages. We curated a representative dataset of over 10,000 proteins derived from NCBI, for which we predicted protein structures with ColabFold and assessed structural similarity via FoldSeek against the PDB, AlphaFold, and Phold databases. We evaluated multiple annotation strategies, including sequence-based (Pharokka), and structure-based (FoldSeek, Phold) methods. Our results show that up to 43 % of truly unannotated proteins can be functionally annotated when combining structure-informed approaches with UniProt-derived annotations. We highlight the complementarity of different databases and the importance of annotation quality filtering. This work provides a valuable resource of predicted structures and annotations, and offers insights into optimizing structure-based annotation pipelines for viral proteins, paving the way for deeper exploration of phage biology and its applications.

Authors 7

1. Hannelore Longin

   KU Leuven

   Affiliation as printed

   Computational Systems Biology, Department of Molecular and Microbial Systems, KU Leuven, Heverlee, Belgium

   Laboratory of Gene Technology, Department of Biosystems, KU Leuven, Heverlee, Belgium

2. George Spyro Bouras

   Basil Hetzel Institute · The University of Adelaide

   Affiliation as printed

   Adelaide Medical School, Faculty of Health and Medical Sciences, The University of Adelaide, Adelaide, Australia

   The Department of Surgery - Otolaryngology Head and Neck Surgery, University of Adelaide and the Basil Hetzel Institute for Translational Health Research, Central Adelaide, Adelaide, Australia

3. Susanna R. Grigson

   Flinders University

   Affiliation as printed

   Flinders Accelerator for Microbiome Exploration, Flinders University, Adelaide, Australia

4. Robert Alan Edwards

   Flinders University

   Affiliation as printed

   Flinders Accelerator for Microbiome Exploration, Flinders University, Adelaide, Australia

5. Hanne Hendrix

   KU Leuven

   Affiliation as printed

   Laboratory of Gene Technology, Department of Biosystems, KU Leuven, Heverlee, Belgium

6. Rob Lavigne

   KU Leuven

   Affiliation as printed

   Laboratory of Gene Technology, Department of Biosystems, KU Leuven, Heverlee, Belgium

7. Vera van Noort corresponding Aachen Institute of Biology, Leiden University

   Leiden University · KU Leuven

   Affiliation as printed

   Computational Systems Biology, Department of Molecular and Microbial Systems, KU Leuven, Heverlee, Belgium

   Institute of Biology, Leiden University, Leiden, The Netherlands