Double-Labeling Method for Visualization and Quantification of Membrane-Associated Proteins in Lactococcus lactis
Abstract
:1. Introduction
2. Results
2.1. Overproduction and Overexpression of Recombinant Homologous Proteins
2.2. Trafficking of Membrane-Associated Proteins in Live Bacterial Cells
2.3. Optimization of Biarsenical Technology for Membrane-Associated Proteins
2.4. Quantification of Overexpressed Proteins
3. Discussion
3.1. Real-Time Detection and Quantification of Protein Expression in L. lactis Cytoplasm and on Cell Surface
3.2. Ellman’s Reagent in Biarsenical Labeling Technology
3.3. Determination of Translational Pathways for Membrane Associated Proteins in L. lactis
4. Materials and Methods
4.1. Genetic Designation and Engineering
4.2. Cell Culture and Plasmid Transformation
4.3. Plasmid Selection, Isolation, and Sequencing
4.4. Cell Induction for Protein Overexpression
4.5. Cell Disruption and Protein Measurement
4.6. Protein Analysis and Purification
4.7. Surface Protein Staining
4.8. Blocking of Surface-Anchored Tetracysteine Tags
4.9. Intracellular Protein Staining
4.10. Data Analysis
5. Conclusions
Supplementary Materials
Author Contributions
Funding
Data Availability Statement
Acknowledgments
Conflicts of Interest
References
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Hoang, M.N.; Peterbauer, C. Double-Labeling Method for Visualization and Quantification of Membrane-Associated Proteins in Lactococcus lactis. Int. J. Mol. Sci. 2023, 24, 10586. https://doi.org/10.3390/ijms241310586
Hoang MN, Peterbauer C. Double-Labeling Method for Visualization and Quantification of Membrane-Associated Proteins in Lactococcus lactis. International Journal of Molecular Sciences. 2023; 24(13):10586. https://doi.org/10.3390/ijms241310586
Chicago/Turabian StyleHoang, Mai Ngoc, and Clemens Peterbauer. 2023. "Double-Labeling Method for Visualization and Quantification of Membrane-Associated Proteins in Lactococcus lactis" International Journal of Molecular Sciences 24, no. 13: 10586. https://doi.org/10.3390/ijms241310586