The purified thermophilic bacterium PS3 F1β10×His-tag is inserted into the F0F1-ATP synthases of chromatophores isolated from photosynthetic bacteria Rhodospirillum rubrum. The studies of biochemical properties ...The purified thermophilic bacterium PS3 F1β10×His-tag is inserted into the F0F1-ATP synthases of chromatophores isolated from photosynthetic bacteria Rhodospirillum rubrum. The studies of biochemical properties of the hybrid chromatophores show that they have both protons-driving capability and photophosphorylation. The fluorescent actin filaments, as a marker of its orientation by video-microscopic experiment, are connected via Maleimido-C3-NTA to the reconstituted β10×His-tag of F0F1-ATP synthases.The clockwise rotation of FoFt-ATP synthases driven by light is observed directly when viewed from the Fo side to Ft. This system should be valuable for further studying the coupling property of F0F1-ATP synthase.展开更多
Escherichia coli 3-Deoxy-D-manno-octulosonate 8-phosphate（KDO8P） synthase catalyzed the condensation reaction between D-arabinose 5-phosphate（A5P） and phosphoenolpyruvate（PEP） to form KDO8P and inorganic phosph...Escherichia coli 3-Deoxy-D-manno-octulosonate 8-phosphate（KDO8P） synthase catalyzed the condensation reaction between D-arabinose 5-phosphate（A5P） and phosphoenolpyruvate（PEP） to form KDO8P and inorganic phosphate（Pi）. The noncovalent tetrameric association ofKDO8P synthase was observed and dissociated in gas phase by means of electrospray ionization mass spectrometry under the very ＂soft＂ conditions. The results indicate that PEP-bound enzyme generated abundant tetrameric species as well as monomeric species at the ＂soft＂ conditions, whereas, the unbound enzyme favored the formation of a dimeric species. The mass spectra of the mixture of the enzyme with one of substrates, PEP, and A5P or one of products, KDO8P and Pi show that the complex of the unbound enzyme with PEP or Pi was prone to the formation of a monomeric species, whereas, that of the unbound enzyme with A5P or KDO8P was similar to the unbound enzyme. The intensity of the dimeric species increased with the increase of temperature at a collision voltage of 10 V. Taken together, the results presented here suggest that mass spectrometry will be a powerful tool to explore subtile conformational changes and/or subunit-subunit interactions of multiprotein assembly induced by ligand-binding and/or the changes of environmental conditions.展开更多
文摘The purified thermophilic bacterium PS3 F1β10×His-tag is inserted into the F0F1-ATP synthases of chromatophores isolated from photosynthetic bacteria Rhodospirillum rubrum. The studies of biochemical properties of the hybrid chromatophores show that they have both protons-driving capability and photophosphorylation. The fluorescent actin filaments, as a marker of its orientation by video-microscopic experiment, are connected via Maleimido-C3-NTA to the reconstituted β10×His-tag of F0F1-ATP synthases.The clockwise rotation of FoFt-ATP synthases driven by light is observed directly when viewed from the Fo side to Ft. This system should be valuable for further studying the coupling property of F0F1-ATP synthase.
基金Supported by National Hi-tech Research and Development Program of China(No.2006AA02Z154)the National Natural Science Foundation of China(No.20675088) and SRF for ROCS, SEM, China
文摘Escherichia coli 3-Deoxy-D-manno-octulosonate 8-phosphate（KDO8P） synthase catalyzed the condensation reaction between D-arabinose 5-phosphate（A5P） and phosphoenolpyruvate（PEP） to form KDO8P and inorganic phosphate（Pi）. The noncovalent tetrameric association ofKDO8P synthase was observed and dissociated in gas phase by means of electrospray ionization mass spectrometry under the very ＂soft＂ conditions. The results indicate that PEP-bound enzyme generated abundant tetrameric species as well as monomeric species at the ＂soft＂ conditions, whereas, the unbound enzyme favored the formation of a dimeric species. The mass spectra of the mixture of the enzyme with one of substrates, PEP, and A5P or one of products, KDO8P and Pi show that the complex of the unbound enzyme with PEP or Pi was prone to the formation of a monomeric species, whereas, that of the unbound enzyme with A5P or KDO8P was similar to the unbound enzyme. The intensity of the dimeric species increased with the increase of temperature at a collision voltage of 10 V. Taken together, the results presented here suggest that mass spectrometry will be a powerful tool to explore subtile conformational changes and/or subunit-subunit interactions of multiprotein assembly induced by ligand-binding and/or the changes of environmental conditions.
