In Vitro and In Vivo Studies on the Structural Organization of Chs3 from Saccharomyces cerevisiae
Abstract
:1. Introduction
2. Results
2.1. Bioinformatic Analyses to Predict Membrane Topology and Three-Dimensional (3D) Structure Of Chs3 Reveals Gatekeeper and Finger Helices
2.2. Protease Protection Assays Suggest a Revised Model on Membrane Topology of Chs3
2.3. Bimolecular Fluorescence Complementation (BiFC) Experiments Demonstrate Di-/Oligomerization of Chs3 In Vivo
2.4. BiFC Experiments Using Truncated Chs3 Versions Reveal the Site of Self-Interaction
2.5. Tracking Chs3 Di-/Oligomerization in Strains Deficient in Chs3 Trafficking
3. Discussion
4. Experimental Procedures
4.1. Chemicals
4.2. Media
4.3. Construction of Plasmids and Strains
4.4. Complementation Assays in Yeast
4.5. α-Factor Synchronization
4.6. Image Acquisition
4.7. Proteinase K Protection Assays
4.8. Sodium Dodecyl Sulfate Polycarylamide Electrophoresis (SDS-PAGE)/Western Blot
4.9. Bioinformatics
5. Conclusions
Supplementary Materials
Acknowledgements
Author Contributions
Conflicts of Interest
References
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Gohlke, S.; Muthukrishnan, S.; Merzendorfer, H. In Vitro and In Vivo Studies on the Structural Organization of Chs3 from Saccharomyces cerevisiae. Int. J. Mol. Sci. 2017, 18, 702. https://doi.org/10.3390/ijms18040702
Gohlke S, Muthukrishnan S, Merzendorfer H. In Vitro and In Vivo Studies on the Structural Organization of Chs3 from Saccharomyces cerevisiae. International Journal of Molecular Sciences. 2017; 18(4):702. https://doi.org/10.3390/ijms18040702
Chicago/Turabian StyleGohlke, Simon, Subbaratnam Muthukrishnan, and Hans Merzendorfer. 2017. "In Vitro and In Vivo Studies on the Structural Organization of Chs3 from Saccharomyces cerevisiae" International Journal of Molecular Sciences 18, no. 4: 702. https://doi.org/10.3390/ijms18040702