The origin of heterogeneity in the red/green cyanobacteriochrome AnPixJg2

Cyanobacteriochromes (CBCRs) are a recently discovered member of the phytochrome superfamily¹. Like phytochromes they bind a linear tetrapyrrole as a chromophore but in contrast their protein scaffold is more compact. CBCRs have a more diverse spectral tuning that covers the entire visible spectrum. There are at least four different sub-families known in CBCRs out of which the red/green CBCRs have a red-absorbing reactant and a green-absorbing photoproduct. We have carried out classical MD simulations of AnPixJg2 in the red-absorbing form. In the simulations we have considered a histidine residue (H322), that is conserved among red/green CBCRs and is critical for the chromophore binding², in two different protonation states. These are the neutral, singly protonated state (SPH model) and the charged, doubly protonated state (DPH model). In the DPH model, PCB is found to be structurally heterogeneous exhibiting two distinct sub-states in contrast to the SPH model where PCB is homogeneous. We carried out umbrella sampling MD simulations to explore the origin of this different behavior. These simulations have revealed that the energy barrier between the two sub-states in the DPH model is lower compared to the SPH model. The symmetry-adapted perturbation theory (SAPT0) calculations show that three structural factors are critical for the transition between the two sub-states: histidine-water, tryptophan flexibility and tyrosine hydrogen bonding. Based on QM/MM calculations of spectroscopic properties⁴, we have shown that the two sub-states cannot be distinguished based on UV/Vis absorption spectra but exhibit distinct features in CD spectra.

(1) Rockwell, N. C.; Ohlendorf, R.; Möglich, A. Proc. Natl. Acad. Sci. USA 2013, 110 (3), 806–807.

(2) Narikawa, R.; Fukushima, Y.; Ishizuka, T.; Itoh, S.; Ikeuchi, M. J. Mol. Biol. 2008, 380 (5), 844–855.

(3) Wiebeler, C.; Gopalakrishna Rao, A.; Gärtner, W.; Schapiro, I. Angew. Chemie Int. Ed. 2019, 58 (7), 1934–1938.