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000890732 1001_ $$0P:(DE-Juel1)161543$$aArinkin, Vladimir$$b0$$eCorresponding author
000890732 245__ $$aStructural determinants underlying the adduct lifetime in the LOV proteins of Pseudomonas putida
000890732 260__ $$aOxford [u.a.]$$bWiley-Blackwell$$c2021
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000890732 520__ $$aThe primary photochemistry is similar among the flavin-bound sensory domains of light–oxygen–voltage (LOV) photoreceptors, where upon blue-light illumination a covalent adduct is formed on the microseconds time scale between the flavin chromophore and a strictly conserved cysteine residue. In contrast, the adduct-state decay kinetics vary from seconds to days or longer. The molecular basis for this variation among structurally conserved LOV domains is not fully understood. Here, we selected PpSB2-LOV, a fast-cycling (τrec 3.5 min, 20 °C) short LOV protein from Pseudomonas putida that shares 67% sequence identity with a slow-cycling (τrec 2467 min, 20 °C) homologous protein PpSB1-LOV. Based on the crystal structure of the PpSB2-LOV in the dark state reported here, we used a comparative approach, in which we combined structure and sequence information with molecular dynamic (MD) simulations to address the mechanistic basis for the vastly different adduct-state lifetimes in the two homologous proteins. MD simulations pointed toward dynamically distinct structural region, which were subsequently targeted by site-directed mutagenesis of PpSB2-LOV, where we introduced single- and multisite substitutions exchanging them with the corresponding residues from PpSB1-LOV. Collectively, the data presented identify key amino acids on the Aβ-Bβ, Eα-Fα loops, and the Fα helix, such as E27 and I66, that play a decisive role in determining the adduct lifetime. Our results additionally suggest a correlation between the solvent accessibility of the chromophore pocket and adduct-state lifetime. The presented results add to our understanding of LOV signaling and will have important implications in tuning the signaling behavior (on/off kinetics) of LOV-based optogenetic tools.
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000890732 7001_ $$0P:(DE-Juel1)131965$$aGranzin, Joachim$$b1
000890732 7001_ $$0P:(DE-Juel1)131482$$aKrauss, Ulrich$$b2
000890732 7001_ $$0P:(DE-Juel1)131457$$aJaeger, Karl-Erich$$b3
000890732 7001_ $$0P:(DE-Juel1)132029$$aWillbold, Dieter$$b4
000890732 7001_ $$0P:(DE-Juel1)131950$$aBatra-Safferling, Renu$$b5$$eCorresponding author
000890732 773__ $$0PERI:(DE-600)2172518-4$$a10.1111/febs.15785$$gp. febs.15785$$n16$$p4955-4972$$tThe FEBS journal$$v288$$x1742-4658$$y2021
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