Proteome driven re-evaluation and functional annotation of the <it>Streptococcus pyogenes </it>SF370 genome

Abstract

Abstract Background The genome data of Streptococcus pyogenes SF370 has been widely used by many researchers and provides a vast array of interesting findings. Nevertheless, approximately 40% of genes remain classified as hypothetical proteins, and several coding sequences (CDSs) have been unrecognized. In this study, we attempted a shotgun proteomic analysis with a six-frame database that was independent of genome annotation. Results Nine proteins encoded by novel ORFs were found by shotgun proteomic analysis, and their specific mRNAs were verified by reverse transcriptional PCR (RT-PCR). We also provided functional annotations for hypothetical genes using proteomic analysis from three different culture conditions that were separated into three fractions: supernatant, soluble, and insoluble. Consequently, we identified 567 proteins on re-evaluation of the proteomic data using an in-house database comprising 1,697 annotated and nine non-annotated CDSs. We provided functional annotations for 126 hypothetical proteins (18.9% out of the 668 hypothetical proteins) based on their cellular fractions and expression profiles under different culture conditions. Conclusions The list of amino acid sequences that were annotated by genome analysis contains outdated information and unrecognized protein-coding sequences. We suggest that the six-frame database derived from actual DNA sequences be used for reliable proteomic analysis. In addition, the experimental evidence from functional proteomic analysis is useful for the re-evaluation of previously sequenced genomes.