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Ultrastructural localization of the maltose-binding protein within the cell envelope of Escherichia coli
Abstract:
Logarithmically growing cells of Escherichia coli were fixed with glutaraldehyde and incubated with antimaltose-binding protein Fab coupled to horseradish peroxide (molecular weight of the complex 80,000). The position of this complex within the cell envelope was determined by reacting with diaminobenzidine-H2O2, staining with osmium tetroxide and processing for thin section electron microscopy. The following observations were made: (i) induction of the maltose-binding protein resulted in swelling and staining of the outer membrane; (ii) the swelling and staining was more prominent in short cells, less prominent or absent in long cells; (iii) rare examples exhibited granular staining in the space between the plasma membrane and the peptidoglycan layer. These stainings were observable mainly in pole caps; (iv) a mutant lacking the receptor for phage lambda showed altered staining pattern. Treatment of glutaraldehyde-fixed cells with EDTA-lysozyme prevented the specific labelling of the maltose-binding protein.
Insights
This study visualizes the maltose-binding protein in Escherichia coli using electron microscopy. Findings reveal its association with the outer membrane, particularly in shorter cells, and its absence in a phage lambda receptor mutant.
Area of Science:
- Microbiology
- Cell Biology
- Bacterial Ultrastructure
Background:
- The maltose-binding protein (MBP) is crucial for nutrient uptake in Escherichia coli.
- Understanding the precise localization of MBP within the bacterial cell envelope is essential for elucidating transport mechanisms.
Purpose of the Study:
- To determine the subcellular localization of the maltose-binding protein in Escherichia coli.
- To investigate the influence of cell morphology and genetic mutations on MBP localization.
Main Methods:
- Escherichia coli cells were fixed with glutaraldehyde and incubated with a horseradish peroxide-conjugated antibody fragment (Fab) against MBP.
- Immunolabeling was visualized using diaminobenzidine-H2O2 and osmium tetroxide staining.
- Subcellular localization was analyzed via thin-section electron microscopy.
Main Results:
- Induction of MBP led to swelling and staining of the outer membrane, more pronounced in shorter cells.
- Granular staining was observed between the plasma and peptidoglycan layers, predominantly in cell pole caps.
- A mutant lacking the phage lambda receptor exhibited an altered staining pattern, and EDTA-lysozyme treatment disrupted specific MBP labeling.
Conclusions:
- The maltose-binding protein is associated with the outer membrane of Escherichia coli, with localization influenced by cell length.
- The study provides insights into the spatial organization of MBP and its potential interaction with other cell envelope components.