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J Photochem Photobiol B. 2014 Nov;140:182-93. doi: 10.1016/j.jphotobiol.2014.06.017. Epub 2014 Aug 7.

Absorption and emission spectroscopic characterization of photo-dynamics of photoactivated adenylyl cyclase mutant bPAC-Y7F of Beggiatoa sp.

Author information

  • 1Fakultät für Physik, Universität Regensburg, Universitätsstrasse 31, D-93053 Regensburg, Germany. Electronic address: alfons.penzkofer@physik.uni-regensburg.de.
  • 2Institut für Biologie/Experimentelle Biophysik, Humboldt Universität zu Berlin, Invalidenstrasse 42, D-10115 Berlin, Germany.
  • 3Institut für Biologie/Experimentelle Biophysik, Humboldt Universität zu Berlin, Invalidenstrasse 42, D-10115 Berlin, Germany; Department of Exact Sciences/Biophysics, Vrije Universiteit, De Boelelaan 1081A, 1081 HV Amsterdam, The Netherlands.

Abstract

The photoactivated cyclase bPAC of the microbial mats bacterium Beggiatoa sp. consists of a BLUF domain and an adenylyl cyclase domain. It has strong activity of photo-induced cyclic adenylyl monophosphate (cAMP) formation and is therefore an important optogenetic tool in neuroscience applications. The SUMO-bPAC-Y7F mutant where Tyr-7 is replaced by Phe-7 in the BLUF domain has lost the typical BLUF domain photo-cycle dynamics. Instead, the investigated SUMO-bPAC-Y7F mutant consisted of three protein conformations with different triplet based photo-dynamics: (i) reversible flavin quinone (Fl) cofactor reduction to flavin semiquinone (FlH), (ii) reversible violet/near ultraviolet absorbing flavin photoproduct (FlA) formation, and (iii) irreversible red absorbing flavin photoproduct (FlC) formation. Absorption and emission spectroscopic measurements on SUMO-bPAC-Y7F were carried out before, during and after light exposure. Flavin photo-dynamics schemes are developed for the SUMO-bPAC-Y7F fractions performing photo-induced FlH, FlA, and FlC formation. Quantitative parameters of the flavin cofactor excitation, relaxation and recovery dynamics in SUMO-bPAC-Y7F are determined.

Copyright © 2014 Elsevier B.V. All rights reserved.

KEYWORDS:

Anionic flavin hydoquinone charge transfer complex formation; BLUF domain; Flavin-C4a-adduct photo-cycle; Photoactivated cyclase bPAC; Soil bacterium Beggiatoa sp.; Triplet based flavin semiquinone photo-cycle

PMID:
25154810
[PubMed - indexed for MEDLINE]
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