Comparative Membrane Proteomics Reveals a Nonannotated E. coli Heat Shock Protein

• Published in: Biochemistry (Easton) Vol. 57; no. 1; pp. 56 - 60
• Main Authors: Yuan, Peijia, D’Lima, Nadia G, Slavoff, Sarah A
• Format: Journal Article
• Language: English
• Published: United States American Chemical Society 09.01.2018
• Subjects: Chromatography, High Pressure Liquid; Communication; Escherichia coli - enzymology; Escherichia coli - physiology; Escherichia coli K12 - enzymology; Escherichia coli K12 - growth & development; Escherichia coli K12 - physiology; Escherichia coli Proteins - chemistry; Escherichia coli Proteins - genetics; Escherichia coli Proteins - metabolism; Gene Expression Regulation, Bacterial; Green Fluorescent Proteins - chemistry; Green Fluorescent Proteins - genetics; Green Fluorescent Proteins - metabolism; Heat-Shock Proteins, Small - chemistry; Heat-Shock Proteins, Small - genetics; Heat-Shock Proteins, Small - metabolism; Heat-Shock Response; Membrane Proteins - chemistry; Membrane Proteins - genetics; Membrane Proteins - metabolism; Models, Molecular; Molecular Sequence Annotation; Open Reading Frames; Phosphogluconate Dehydrogenase - chemistry; Phosphogluconate Dehydrogenase - genetics; Phosphogluconate Dehydrogenase - metabolism; Protein Conformation, alpha-Helical; Protein Interaction Domains and Motifs; Protein Transport; Proteogenomics - methods; Proteomics - methods; Recombinant Fusion Proteins - chemistry; Recombinant Fusion Proteins - metabolism; Tandem Mass Spectrometry
• ISSN: 0006-2960; 1520-4995
• DOI: 10.1021/acs.biochem.7b00864

Recent advances in proteomics and genomics have enabled discovery of thousands of previously nonannotated small open reading frames (smORFs) in genomes across evolutionary space. Furthermore, quantitative mass spectrometry has recently been applied to analysis of regulated smORF expression. However, bottom-up proteomics has remained relatively insensitive to membrane proteins, suggesting they may have been underdetected in previous studies. In this report, we add biochemical membrane protein enrichment to our previously developed label-free quantitative proteomics protocol, revealing a never-before-identified heat shock protein in Escherichia coli K12. This putative smORF-encoded heat shock protein, GndA, is likely to be ∼36–55 amino acids in length and contains a predicted transmembrane helix. We validate heat shock-regulated expression of the gndA smORF and demonstrate that a GndA-GFP fusion protein cofractionates with the cell membrane. Quantitative membrane proteomics therefore has the ability to reveal nonannotated small proteins that may play roles in bacterial stress responses.