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The characterization of mannan of Micrococcus lysodeikticus as an acidic lipopolysaccharide
Abstract:
Ghosts of Micrococcus lysodeikticus contain a mannan that is not removed by intensive washing procedures. Purified mannan, isolated by extraction of whole cells with hot, aqueous phenol, binds to membranes in vitro. Mannan also binds to DEAE-cellulose and migrates toward the anode in neutral and sodium dodecyl sulfate disc gel electrophoresis. In aqueous solution mannan has an apparent molecular weight of 10-6, but in the presence of sodium dodecyl sulfate its apparent molecular weight is 50,000 to 100,000; removal of the detergent results in reaggregation. Purified mannan contains mannose, succinate, fatty acid, and glycerol in a ratio of 50:4.9:2.1:1.0. Treatment of mannan with mild base produces a neutral, hydrophilic polysaccharide of relatively low molecular weight that has no affinity for membranes. At least 90% of the reducing termini are blocked in a base-stable linkage. Based on these results a tentative structure is proposed for the mannan.
Insights
Micrococcus lysodeikticus ghosts contain a resilient mannan. This polysaccharide binds to membranes and DEAE-cellulose, with a proposed structure based on its composition and behavior.
Area of Science:
- Microbiology
- Biochemistry
- Structural Biology
Background:
- Micrococcus lysodeikticus cell envelopes contain a mannan component.
- This mannan is resistant to extensive washing procedures, suggesting strong association with cellular structures.
Purpose of the Study:
- To isolate and characterize the mannan from Micrococcus lysodeikticus ghosts.
- To elucidate the binding properties and structural features of this bacterial mannan.
Main Methods:
- Extraction of whole cells using hot, aqueous phenol to purify mannan.
- In vitro binding assays with membranes and DEAE-cellulose.
- Electrophoresis (neutral and SDS-PAGE) to determine charge and molecular weight.
- Chemical analysis to identify constituent components and their ratios.
Main Results:
- Purified mannan exhibits binding affinity for both biological membranes and DEAE-cellulose.
- Electrophoretic analysis indicates mannan carries a net negative charge and aggregates in SDS, with molecular weights ranging from 50,000-100,000.
- Composition analysis reveals mannose, succinate, fatty acid, and glycerol in a specific ratio.
- Mild base treatment yields a non-binding, low molecular weight polysaccharide, indicating base-labile linkages.
Conclusions:
- The Micrococcus lysodeikticus mannan is a complex polysaccharide with significant membrane-binding capabilities.
- A tentative structure is proposed, highlighting the presence of blocked reducing termini in base-stable linkages.