Abstract
Far-red absorbing allophycocyanins (APC), identified in cyanobacteria capable of FRL photoacclimation (FaRLiP) and low-light photoacclimation (LoLiP), absorb far-red light, functioning in energy transfer as light-harvesting proteins. We report an optimized method to obtain high purity far-red absorbing allophycocyanin B, AP-B2, of Chroococcidiopsis thermalis sp. PCC7203 by synthesis in Escherichia coli and an improved purification protocol. The crystal structure of the trimer, (PCB-ApcD5/PCB-ApcB2)3, has been resolved to 2.8 Å. The main difference to conventional APCs absorbing in the 650–670 nm range is a largely flat chromophore with the co-planarity extending, in particular, from rings BCD to ring A. This effectively extends the conjugation system of PCB and contributes to the super-red-shifted absorption of the α-subunit (λmax = 697 nm). On complexation with the β-subunit, it is even further red-shifted (λmax, absorption = 707 nm, λmax, emission = 721 nm). The relevance of ring A for this shift is supported by mutagenesis data. A variant of the α-subunit, I123M, has been generated that shows an intense FR-band already in the absence of the β-subunit, a possible model is discussed. Two additional mechanisms are known to red-shift the chromophore spectrum: lactam-lactim tautomerism and deprotonation of the chromophore that both mechanisms appear inconsistent with our data, leaving this question unresolved.
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Data availability
All data that are relevant for our conclusions are present in this article and/or the Supplementary Materials. The atomic coordinate file of the AP-B2 structure has been deposited in the Protein Data Bank with accession code 8OR7.
Notes
Strictly speaking, this takes the line between C4 and C6 as a pseudo-bond.
Abbreviations
- AP-B:
-
Allophycocyanin-B
- APC:
-
Allophycocyanin
- FRL:
-
Far-red light
- FaRLiP:
-
Far-red light photoacclimation
- LoLiP:
-
Low-light photoacclimation
- PBP:
-
Phycobiliprotein
- PBS:
-
Phycobilisome
- PCB:
-
Phycocyanobilin
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Acknowledgements
We thank Hong Zhang at Huazhong University of Science and Technology of China for helping with X-ray data collection. We thank the Shanghai Synchrotron Radiation Facility (SSRF) BL18U for beamline support.
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This work was supported by the National Natural Science Foundation of China (Grant 31861143029 to K.H.Z). The Center for Structural Studies is funded by the Deutsche Forschungsgemeinschaft (DFG Grant Number 417919780 and INST 208/740-1 FUGG).
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L.J.Z., Y.Q.W., and J.Y.H. performed investigations. L.J.Z., A.H., H.S, and K.H.Z. analyzed data. L.J.Z., A.H., H.S, and K.H.Z. wrote the main manuscript text. All authors reviewed the manuscript.
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Zhou, LJ., Höppner, A., Wang, YQ. et al. Crystallographic and biochemical analyses of a far-red allophycocyanin to address the mechanism of the super-red-shift. Photosynth Res (2024). https://doi.org/10.1007/s11120-023-01066-2
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DOI: https://doi.org/10.1007/s11120-023-01066-2