ABSTRACT
Small proteins (SPs) are defined as peptides of 100 amino acids or less encoded by short open reading frames (sORFs). SPs participate in a wide range of functions in cells, including gene regulating, cell signaling and metabolism. However, most annotated SPs in all living organisms are currently lacking expression evidence at the protein level and regarded as missing proteins (MPs). High efficient SPs identification is the prerequisite for their functional study and contribution to MPs searching. In this study, we identified 72 SPs and successfully validated 9 MPs from Saccharomyces cerevisiae based on SPs enrichment strategy. In-depth analysis showed that the missing factors of MPs were low molecular weight, low abundant, hydrophobicity, lower codon usage bias and unstable. The small protein-based enrichment can be used as MPs searching strategy, which might provide the foundation for their further function research.