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Characterization of chlorophyll f synthase heterologously produced in Synechococcus sp. PCC 7002

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Abstract

In diverse terrestrial cyanobacteria, Far-Red Light Photoacclimation (FaRLiP) promotes extensive remodeling of the photosynthetic apparatus, including photosystems (PS)I and PSII and the cores of phycobilisomes, and is accompanied by the concomitant biosynthesis of chlorophyll (Chl) d and Chl f. Chl f synthase, encoded by chlF, is a highly divergent paralog of psbA; heterologous expression of chlF from Chlorogloeopsis fritscii PCC 9212 led to the light-dependent production of Chl f in Synechococcus sp. PCC 7002 (Ho et al., Science 353, aaf9178 (2016)). In the studies reported here, expression of the chlF gene from Fischerella thermalis PCC 7521 in the heterologous system led to enhanced synthesis of Chl f. N-terminally [His]10-tagged ChlF7521 was purified and identified by immunoblotting and tryptic-peptide mass fingerprinting. As predicted from its sequence similarity to PsbA, ChlF bound Chl a and pheophytin a at a ratio of ~ 3–4:1, bound β-carotene and zeaxanthin, and was inhibited in vivo by 3-(3,4-dichlorophenyl)-1,1-dimethylurea. Cross-linking studies and the absence of copurifying proteins indicated that ChlF forms homodimers. Flash photolysis of ChlF produced a Chl a triplet that decayed with a lifetime (1/e) of ~ 817 µs and that could be attributed to intersystem crossing by EPR spectroscopy at 90 K. When the chlF7521 gene was expressed in a strain in which the psbD1 and psbD2 genes had been deleted, significantly more Chl f was produced, and Chl f levels could be further enhanced by specific growth-light conditions. Chl f synthesized in Synechococcus sp. PCC 7002 was inserted into trimeric PSI complexes.

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Acknowledgements

This work was supported by the National Science Foundation grant MCB-1613022 to D.A.B and J.H.G. This research was also conducted under the auspices of the Photosynthetic Antenna Research Center (PARC), an Energy Frontier Research Center funded by the DOE, Office of Science, Office of Basic Energy Sciences under Award Number DE-SC 0001035 (D.A.B.). A.v.d.E. acknowledges support from the Natural Science and Engineering Research Council, Canada in the form of a Discovery Grant. D.P.C. was supported by a European Commission Marie Skłodowska-Curie Global Fellowship (660652). The authors thank Yue Lu at Washington University in St. Louis for performing the mass spectrometric analyses.

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  1. Daniel P. Canniffe

    Present address: Department of Molecular Biology & Biotechnology, University of Sheffield, Sheffield, S10 2TN, UK

Authors and Affiliations

  1. Department of Biochemistry and Molecular Biology, The Pennsylvania State University, University Park, PA, 16802, USA

    Gaozhong Shen, Daniel P. Canniffe, Ming-Yang Ho, Vasily Kurashov, John H. Golbeck & Donald A. Bryant

  2. Department of Chemistry, The Pennsylvania State University, University Park, PA, 16802, USA

    John H. Golbeck

  3. Department of Chemistry, Brock University, St. Catharines, ON, L2S 3A1, Canada

    Art van der Est

  4. Department of Chemistry and Biochemistry, Montana State University, Bozeman, MT, 59717, USA

    Donald A. Bryant

  5. S-002 Frear Laboratory, Department of Biochemistry and Molecular Biology, The Pennsylvania State University, University Park, PA, 16802, USA

    Donald A. Bryant

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Shen, G., Canniffe, D.P., Ho, MY. et al. Characterization of chlorophyll f synthase heterologously produced in Synechococcus sp. PCC 7002. Photosynth Res 140, 77–92 (2019). https://doi.org/10.1007/s11120-018-00610-9

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