Searching sequence databases for functional homologs using profile HMMs: how to set bit score thresholds?

ABSTRACT Motivation UniProt and BFD databases together have 2.5 billion protein sequences. A large majority of these proteins have been electronically annotated. Automated annotation pipelines, vis-à-vis manual curation, have the advantage of scale and speed but are fraught with relatively higher error rates. This is because sequence homology does not necessarily translate to functional homology, molecular function specification is hierarchic and not all functional families have the same amount of experimental data that one can exploit for annotation. Consequently, customization of annotation workflow is inevitable to minimize annotation errors. Results We discuss possible ways of customizing the search of sequence databases for functional homologs using profile HMMs. Choosing an optimal bit score threshold is a critical step in the application of HMMs, which is illustrated using four Case Studies; the single domain nucleotide sugar 6-dehydrogenase and lysozyme-C families, and SH3 and GT-A domains which are typically found as a part of multi-domain proteins. We also discuss the limitations of using profile HMMs for functional annotation and suggests some possible ways to partially overcome such limitations. Supplementary information Supplementary_material containing Figures S1-S7 and Tables S1 and S2 Supplementary_dataset.xlsx