Colicin-Bearing Plasmids Carried by Shiga Toxin-Producing E. coli (STEC) Analyzed by Targeted Top-Down MS Analysis
Clifton K Fagerquist1, Yanlin Shi1, Mahesh Koirala1,2
1Produce Safety and Microbiology, Western Regional Research Center, Agricultural Research Service, US Department of Agriculture, Albany, California, USA.
Rapid Communications in Mass Spectrometry : RCM
|January 24, 2026
Summary
This study identifies colicin immunity proteins in Shiga toxin-producing E. coli using mass spectrometry. This method rapidly detects antibacterial toxins on bacterial plasmids, aiding pathogen survival research.
Area of Science:
- Microbiology
- Bacterial Genetics
- Proteomics
Background:
- Colicins are bacterial protein toxins targeting nucleic acids or membranes, encoded on plasmids crucial for bacterial survival.
- Identifying molecular factors aiding pathogen survival is vital, even if colicins don't directly cause virulence.
Purpose of the Study:
- To develop and validate a rapid method for detecting colicin-bearing plasmids in Shiga toxin-producing E. coli (STEC).
- To identify colicin immunity proteins and analyze their genetic regulation in STEC strains.
Main Methods:
- Analysis of STEC strains using antibiotic induction, MALDI-TOF-TOF, Orbitrap mass spectrometry, and targeted top-down analysis.
- Small plasmid sequencing to confirm colicin/immunity genes and their upstream regulation.
- Utilizing AlphaFold3 to interpret mass spectrometry fragmentation patterns.
Main Results:
- Detection and identification of colicin immunity proteins (ImBac, ImmD, and a hypothetical protein) in two STEC strains (O26:H11, O104:H7).
- Confirmation of colicin and immunity genes and their regulatory elements through plasmid sequencing.
- MALDI-TOF-TOF-MS/MS-PSD demonstrated effectiveness for identifying inducible antibacterial toxins.
Conclusions:
- MALDI-TOF-TOF-MS/MS-PSD is a powerful platform for rapid detection of bacterial protein toxins.
- The glycine-enhanced aspartic acid effect (AAE) is frequently observed in unstructured protein regions.
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