Sang-Hun Song, Peter Freddolino, Abigail Nash, Elizabeth Carroll, Klaus
Schulten, Kevin Gardner, and Delmar S. Larsen.
Modulating LOV domain photodynamics with a residue alteration
outside the chromophore binding site.
Biochemistry, 50:2411-2423, 2011.
(PMC: 3068209)
SONG2011
Phototropins, a class of light-activated protein kinases, are essential for several blue light
responses in plants and algae, including phototropism. These proteins contain two
internal light, oxygen, and voltage sensitive (LOV) domains, which bind flavin
chromophores and undergo a reversible photochemical formation of a cysteinyl-flavin
adduct as part of the light sensing process. While the photodynamic properties of such
photosensory domains are dictated by interactions between the chromophore and
surrounding protein, more distant residues can play a significant role as well. Here we
explore the role of the Phe434 residue in the photosensory response of the second LOV
domain of Avena sativa phototropin 1 (AsLOV2), a model photochemical system for
these LOV domains. Phe434 lies over 6 Åfrom the FMN chromophore in AsLOV2;
nevertheless, a F434Y point mutation is likely to change several structural features of the
chromophore binding site, as we demonstrate using molecular dynamics simulations.
Transient absorption signals spanning 15 decades in time were compared for wildtype
AsLOV2 and the F434Y mutant, showing that the latter has significantly altered
photodynamics including (i) a faster intersystem crossing leading to triplet formation on a
nanosecond time scale, (ii) biphasic formation of adduct state kinetics on the microsecond
time scale, and (iii) greatly accelerated ground-state recovery kinetics on a second time
scale. We present mechanistic models that link these spectroscopic differences to changes
in the configuration of the critical cysteine residue and in the chromophore's accessibility
to solvent and oxygen according to MD trajectories and purging experiments. Taken
together, these results demonstrate the importance of residues outside the chromophore-
binding pocket in modulating LOV domain photodynamics.
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