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Site-specific protein dynamics in communication pathway from sensor to signaling domain of oxygen sensor protein, HemAT-Bs: Time-resolved Ultraviolet Resonance Raman Study
Authors:El-Mashtoly Samir F  Kubo Minoru  Gu Yuzong  Sawai Hitomi  Nakashima Satoru  Ogura Takashi  Aono Shigetoshi  Kitagawa Teizo
Affiliation:From the Lehrstuhl für Biophysik, Ruhr-Universität Bochum, 44780 Bochum, Germany.;the §Picobiology Institute, Graduate School of Life Science, University of Hyogo, Hyogo 678-1297, Japan, and ;the Okazaki Institute for Integrative Bioscience, Okazaki 444-8787, Japan
Abstract:HemAT-Bs is a heme-based signal transducer protein responsible for aerotaxis. Time-resolved ultraviolet resonance Raman (UVRR) studies of wild-type and Y70F mutant of the full-length HemAT-Bs and the truncated sensor domain were performed to determine the site-specific protein dynamics following carbon monoxide (CO) photodissociation. The UVRR spectra indicated two phases of intensity changes for Trp, Tyr, and Phe bands of both full-length and sensor domain proteins. The W16 and W3 Raman bands of Trp, the F8a band of Phe, and the Y8a band of Tyr increased in intensity at hundreds of nanoseconds after CO photodissociation, and this was followed by recovery in ~50 μs. These changes were assigned to Trp-132 (G-helix), Tyr-70 (B-helix), and Phe-69 (B-helix) and/or Phe-137 (G-helix), suggesting that the change in the heme structure drives the displacement of B- and G-helices. The UVRR difference spectra of the sensor domain displayed a positive peak for amide I in hundreds of nanoseconds after photolysis, which was followed by recovery in ~50 μs. This difference band was absent in the spectra of the full-length protein, suggesting that the isolated sensor domain undergoes conformational changes of the protein backbone upon CO photolysis and that the changes are restrained by the signaling domain. The time-resolved difference spectrum at 200 μs exhibited a pattern similar to that of the static (reduced - CO) difference spectrum, although the peak intensities were much weaker. Thus, the rearrangements of the protein moiety toward the equilibrium ligand-free structure occur in a time range of hundreds of microseconds.
Keywords:Heme   Raman Spectroscopy   Signal Transduction   Spectroscopy   UV Spectroscopy   Gas Sensor Proteins   Oxygen Sensor   Protein Dynamics   Time-resolved Resonance Raman
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