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| Effect of D97N Mutation on Proton Transport and Energy Conversion in the Photoreceptor Archaerhodopsin 4 |
| WANG Juan, GAO Yu-jiao, SUN Chao, ZHAO Xin |
| Shanghai Key Laboratory of Magnetic Resonance, School of Physics and Materials Science, East China Normal University, Shanghai 200062, China |
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Abstract Archaerhodopsin 4 (aR4) is a newly discovered photosensitive retinal protein from the claret membrane of Halobacterium species xz515. It functions as a proton pump similar to bacteriorhodopsin (bR) but with an opposite temporal order of proton uptake and release at neutral pH. Aspartic acid 97 (D97) is one of the key residues as the proton donor during the photocycle in aR4. Its function on the photocycle, the proton pump function and the energy conversion is very important for a deep understanding of the relationships between aR4 structure and function, especially with proton release and uptake order. Single mutation of D96N of bR and D97N of aR4 have been successfully constructed by combining site-directed mutagenesis with heterologous expression in Halobacterium species L33. The influences of mutations on the retinal binding pocket, transient kinetic change of the M state and O state, and the ATP conversion efficiency have been studied by UV-VIS and flashlight induced kinetic absorption change spectroscopy. The results show that D97N mutation in aR4 does not affect the UV absorption of the retinal chromophore, but results in a significant prolongation of the M state, a greatly weakened proton pump, and a decreased ATP formation efficiency. This indicates a different function of D97 in aR4 than the same type residue D96 in bR due to a more hydrophobic environment along the proton translocation channel from D97 to the retinal chromophore.
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Received: 21 February 2017
Published: 25 September 2017
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