Project description:Short open reading frames (sORF) have the potential to encode small proteins in the human gut microbiome, but their role in this environment is rarely understood. These small proteins, known as sORF-encoded peptides (SEP), are less than or equal to 100 amino acids in length. Using bottom-up proteomics (BUP) and top-down proteomics (TDP) approaches, we aimed to identify SEP under various growth conditions and stress exposure in Blautia producta, a key species in mediating colonization resistance and dampening gut inflammation. After applying a rigorous filtering and validation procedure, we identified 45 SEP. Our findings indicate that in contrast to previous results the production of specific SEP is not restricted to direct bacterial interactions within the microbiome but rather depends on growth and stress conditions of the environment. Top-down analysis improved identification confidence and detected a number of full-length with and without N-terminal methionine excision (NME), N- or C-terminal truncated or post-translational modified proteoforms of SEP, indicating that these are common occurrences in B. producta. These findings highlight SEP as a ubiquitous class of non-annotated polypeptides that have been overlooked as a subset of proteins in B. producta.
