Abstract
Biliproteins are a unique class of photosynthetic proteins in their diverse, and at times, divergent biophysical function. The two contexts of photosynthetic light harvesting and photoreception demonstrate characteristically opposite criteria for success, with light harvesting demanding structurally-rigid chromophores which minimize excitation quenching, and photoreception requiring structural flexibility to enable conformational isomerization. The functional plasticity borne out in these two biological contexts is a consequence of the structural plasticity of the pigments utilized by biliproteins―linear tetrapyrroles, or bilins. In this work, the intrinsic flexibility of the bilin framework is investigated in a bottom-up fashion by reducing the active nuclear degrees of freedom through model dipyrrole subunits of the bilin core and terminus free of external protein interactions. Steady-state spectroscopy was carried out on the dipyrrole (DPY) and dipyrrinone (DPN) subunits free in solution to characterize their intrinsic spectroscopic properties including absorption strengths and nonradiative activity. Transient absorption (TA) spectroscopy was utilized to determine the mechanism and kinetics of nonradiative decay of the dipyrrole subunits, revealing dynamics dominated by rapid internal conversion with some Z→E isomerization observable in DPY. Computational analysis of the ground state conformational landscapes indicates enhanced complexity in the asymmetric terminal subunit, and the prediction was confirmed by heterogeneity of species and kinetics observed in TA. Taken together, the large oscillator strengths (f ∼ 0.6) of the dipyrrolic derivatives and chemically-efficient spectral tunability seen through the ∼100 nm difference in absorption spectra, validate Nature's "selection" of multi-pyrrole pigments for light capture applications. However, the rapid deactivation of the excited state via their natural torsional activity when free in solution would limit their effective biological function. Comparison with phytochrome and phycocyanin 645 crystal structures reveals binding motifs within the in vivo bilin environment that help to facilitate or inhibit specific inter-pyrrole twisting vital for protein operation.
Highlights
The light harvesting process in photosynthesis constitutes the initial step that triggers the subsequent chain of electron transfer events and chemical reactions
Studies on single dipyrrinone derivatives have yielded similar results, with rapid deactivation attributed to twisting motion of the two rings leading mainly to the recovery of the ground state Z isomer (Lamola et al, 1983; Zietz and Gillbro, 2007; Sakata et al, 2016). These results suggest the intrinsic nature of the dipyrrinone bilin subunit to undergo ring twisting in the excited state when free in solution, and a seminal work by Lightner et al elegantly demonstrated the suppression of this behavior by chemically linking the two rings
The torsional freedom of linear tetrapyrroles provides both spectral tunability when external contacts are made to contort the local geometry across a dipyrrole pair, or the ability to isomerize across the methine bridge of a dipyrrole pair
Summary
The light harvesting process in photosynthesis constitutes the initial step that triggers the subsequent chain of electron transfer events and chemical reactions. It involves first the absorption of light by a chromophore, or pigment, followed by the transport of that energy to a reaction center site where it is converted into a charge separation This process typically occurs at efficiencies approaching unity, despite the tens to hundreds of pigment-pigment energy transfer events required to span the spatial extent of the light-harvesting apparatus (Glazer, 1989; MacColl, 1998; Wientjes et al, 2013; Mirkovic et al, 2017). Such remarkable efficiencies are accomplished by utilization of antenna proteins that augment reaction centers for dramatically enhanced light capture, while simultaneously providing a pigment-coupling network to facilitate rapid and directional excitation transport prior to quenching processes. Among the various cryptophyte phycobiliproteins sequenced and whose crystal structures have been solved, significant homology is demonstrated both in sequence and αβ conformation (Harrop et al, 2014), showing that light capture differences occurs predominantly by exchanging bilin types
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