Characterization of the Cph1 holo-phytochrome from Synechocystis sp. PCC 6803.
The cph1 gene from the unicellular cyanobacterium Synechoycstis sp. PCC 6803 encodes a protein with the characteristics of plant phytochromes and histidine kinases of two-component phospho-relay systems. Spectral and biochemical properties of Cph1 have been intensely studied in vitro using protein from recombinant systems, but virtually nothing is known about the situation in the natural host. In the present study, His6-tagged Cph1 was isolated from Synechocystis cells. The cph1-his gene was expressed either under the control of the natural cph1 promoter or over-expressed using the strong promoter of the psbA2 gene. Upon purification with nickel affinity chromatography, the presence of Cph1 in extracts was confirmed by immunoblotting and Zn2+-induced fluorescence. The Cph1 extracts exhibited a red/far-red photoactivity characteristic of phytochromes. Difference spectra were identical with those of the phycocyanobilin adduct of recombinant Cph1, implying that phycocyanobilin is the chromophore of Cph1 in Synechocystis.
Humboldt Universität Berlin, Institut für Biologie/Genetik, Berlin, Germany. thomas=huebschmann@biologie.hu-berlin.de
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D001426:Bacterial Proteins; D015152:Blotting, Northern; D002846:Chromatography, Affinity; D000458:Cyanobacteria / Q000201:enzymology* / Q000235:genetics; D004591:Electrophoresis, Polyacrylamide Gel; D005453:Fluorescence; D006639:Histidine / Q000235:genetics / Q000378:metabolism; D009532:Nickel; D020283:Photoreceptors, Microbial; D052980:Phycobilins; D010798:Phycocyanin / Q000737:chemistry; D010834:Phytochrome / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification*; D011401:Promoter Regions, Genetic; D011494:Protein Kinases / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification*; D011758:Pyrroles / Q000737:chemistry; D011994:Recombinant Proteins / Q000737:chemistry; D013054:Spectrophotometry, Atomic; D045725:Tetrapyrroles; D014158:Transcription, Genetic; D015032:Zinc / Q000378:metabolism
D001426:Bacterial Proteins; D020283:Photoreceptors, Microbial; D052980:Phycobilins; D011758:Pyrroles; D011994:Recombinant Proteins; D045725:Tetrapyrroles; D010798:Phycocyanin; D010834:Phytochrome; C011069:phycocyanobilin; D006639:Histidine; D009532:Nickel; D011494:Protein Kinases; C413623:Cph1 phytochrome protein, bacteria; D015032:Zinc