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Mannose incorporation and lectin recognition of pronase-sensitive components in Micrococcus lysodeikticus (M. luteus)
Abstract:
Isolated cytoplasmic membranes from Micrococcus lysodeikticus were able to incorporate [14C]mannose from GDP-[14C]mannose. Labelled mannose remained in the membrane fraction after its repeated washing and lipid extraction. Sodium dodecyl sulfate gel electrophoresis in 12% acrylamide showed a set of bands with molecular weights ranging from 230 000 to 19 000 which stained for protein and carbohydrate, and incorporated [14C]mannose. Some of these bands reacted with different lectins (concanavalin A, wheat germ agglutinin and ricin). Furthermore, the mannose was incorporated via a glycosylation pathway similar to that followed in eukaryotic system as shown by the preliminary identification of a lipid intermediate transferring the sugar to proteins and by the differential sensitivity to bacitracin and tunicamycin. These complex membrane components were sensitive to digestion with pronase. All the results presented suggest their glycoprotein nature.
Insights
Micrococcus lysodeikticus membranes incorporate mannose into glycoproteins via a pathway resembling eukaryotic systems. This study identifies complex membrane glycoproteins involved in mannose incorporation.
Area of Science:
- Microbiology and Biochemistry
- Cellular and Molecular Biology
Background:
- Understanding bacterial membrane composition and function is crucial.
- Glycosylation pathways in prokaryotes are less understood compared to eukaryotes.
Purpose of the Study:
- To investigate the incorporation of mannose into membrane components of Micrococcus lysodeikticus.
- To characterize the nature of these mannose-incorporating membrane molecules.
Main Methods:
- Isolation of cytoplasmic membranes from Micrococcus lysodeikticus.
- Incorporation of GDP-[14C]mannose and subsequent analysis.
- Sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) and lectin binding assays.
- Sensitivity to pronase digestion and analysis of glycosylation pathway inhibitors.
Main Results:
- Micrococcus lysodeikticus membranes incorporated [14C]mannose from GDP-[14C]mannose.
- SDS-PAGE revealed protein and carbohydrate-staining bands that incorporated mannose, with molecular weights from 19,000 to 230,000 Da.
- Some bands showed reactivity with lectins (concanavalin A, wheat germ agglutinin, ricin), and mannose incorporation followed a pathway similar to eukaryotes, involving a lipid intermediate and sensitivity to bacitracin and tunicamycin.
Conclusions:
- The results strongly suggest the presence of glycoproteins in Micrococcus lysodeikticus membranes.
- The identified glycosylation pathway shares similarities with eukaryotic systems, indicating conserved mechanisms.
- These complex membrane components are likely involved in essential cellular processes.