Pseudomonas fluorescens proliferates in a mouse organ homogenate at low temperature
Yota Tatara1, Takahiro Terakawa, Youhei Yamagata
1Laboratory of Enzymology, Graduate School of Agricultural Science, Tohoku University, Sendai, Miyagi, Japan.
Abstract:
In this study we observed the proliferation of Pseudomonas fluorescens (P. fluorescens) in mouse organ homogenates at 4 degrees C. P. fluorescens secreted a protease possessing properties different from those of the mammalian tissue proteases. The specificity of this protease required a basic amino acid residue at the P1 position at a pH optimum of 6.0. The specificity of the protease was similar to that of trypsin, but the pH optimum was different. The protease mildly degraded elastin-Congo red; this suggests that the protease serves as an alternative for elastase in the case of P. fluorescens strains that lack virulent elastase. The protease was identified as an alkaline protease of P. fluorescens by liquid chromatography-tandem mass spectrometry analysis. Our results show that proteome analysis of the soluble proteins is useful in identifying bacterial species, particularly the bacterial contaminants in samples containing antibiotics.
Insights
Pseudomonas fluorescens secretes a novel alkaline protease that degrades elastin, potentially acting as an elastase alternative. Proteome analysis aids in identifying bacterial contaminants in antibiotic-containing samples.
Area of Science:
- Microbiology
- Biochemistry
- Proteomics
Background:
- Pseudomonas fluorescens (P. fluorescens) can proliferate in biological samples.
- Bacterial proteases can degrade host tissues and interfere with sample analysis.
- Identifying bacterial contaminants is crucial, especially in samples containing antibiotics.
Purpose of the Study:
- To characterize a protease secreted by P. fluorescens.
- To investigate the potential role of this protease as an elastase alternative.
- To evaluate the utility of proteome analysis for identifying bacterial contaminants.
Main Methods:
- Observation of P. fluorescens proliferation in mouse organ homogenates.
- Biochemical characterization of the secreted protease's specificity and pH optimum.
- Degradation assay using elastin-Congo red.
- Liquid chromatography-tandem mass spectrometry (LC-MS/MS) for protease identification.
Main Results:
- P. fluorescens secretes a protease with unique properties distinct from mammalian proteases.
- The protease exhibits specificity for basic amino acid residues at the P1 position and a pH optimum of 6.0.
- Mild degradation of elastin-Congo red was observed, suggesting elastase-like activity.
- LC-MS/MS analysis identified the enzyme as an alkaline protease of P. fluorescens.
- Proteome analysis proved effective for identifying bacterial species, including contaminants in antibiotic-treated samples.
Conclusions:
- P. fluorescens produces an alkaline protease with potential elastase-like activity.
- This protease may compensate for the lack of virulent elastase in certain P. fluorescens strains.
- Proteome analysis of soluble proteins is a valuable method for bacterial identification and contaminant detection in complex samples.
