Abstract
Chl. tepidum is a Gram-negative green-sulfur bacterium, which is strict by anaerobic and grows by utilizing sulfide or thiosulfate as an electron source. Blue native-polyacrylamide gel electrophoresis (BN-PAGE) is widely used for the analysis of oligomeric state and molecular mass non-dissociated protein complexes. In this study, a number of proteomic techniques were used to investigate the oligomeric state enzymes. In particular, the Chl. tepidum-soluble proteome was monitored under native condition by using BN-PAGE. The BN-PAGE protein complexes map was analyzed by MALDI-TOF MS after trypsin treatment and from 42 BN proteins bands, 62 different proteins were identified. Additionally, functional information regarding protein–protein interactions was assembled, by coupling 2-D BN-PAGE with MALDI-TOF MS. One-hundred and seventy gel bands were spotted, out of which 187 different proteins were identified. The identified proteins belong to various functional categories like energy metabolism, protein synthesis, amino acid biosynthesis, central intermediate metabolism, and biosynthesis of cofactors indicating the potential of the method for elucidation of functional proteomes.
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Abbreviations
- BN:
-
blue native
- MALDI-TOF:
-
matrix assisted laser desorption ionization – time of flight
- MS:
-
mass spectrometry
- PAGE:
-
polyacrylamide gel electrophoresis
- PMF:
-
peptide mass fingerprinting
- SDS:
-
sodium dodecyl sulfate
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Acknowledgements
The authors thank K. Kouyianou for the critical reading of the manuscript. The project was supported by the University of Crete, the Greek Ministry of Education, and the Centre of Membrane Proteomics (CMP) in Frankfurt.
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Aivaliotis, M., Karas, M. & Tsiotis, G. High throughput two-dimensional blue-native electrophoresis: a tool for functional proteomics of cytoplasmatic protein complexes from Chlorobium tepidum . Photosynth Res 88, 143–157 (2006). https://doi.org/10.1007/s11120-006-9046-z
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DOI: https://doi.org/10.1007/s11120-006-9046-z