Expression of recombinant full-length plant phytochromes assembled with phytochromobilin in Pichia pastoris |
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Authors: | Ah-Young Shin Yun-Jeong Han Pill-Soon Song Jeong-Il Kim |
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Institution: | 1. Department of Biotechnology and Kumho Life Science Laboratory, Chonnam National University, Gwangju 500-757, Republic of Korea;2. Faculty of Biotechnology and Subtropical Horticulture Research Institute, Jeju National University, Jeju 690-756, Republic of Korea |
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Abstract: | We have successfully developed a system to produce full-length plant phytochrome assembled with phytochromobilin in Pichia pastoris by co-expressing apophytochromes and chromophore biosynthetic genes, heme oxygenase (HY1) and phytochromobilin synthase (HY2) from Arabidopsis. Affinity-purified phytochrome proteins from Pichia cells displayed zinc fluorescence indicating chromophore attachment. Spectroscopic analyses showed absorbance maximum peaks identical to in vitro reconstituted phytochromobilin-assembled phytochromes, suggesting that the co-expression system is effective to generate holo-phytochromes. Moreover, mitochondria localization of the phytochromobilin biosynthetic genes increased the efficiency of holophytochrome biosynthesis. Therefore, this system provides an excellent source of holophytochromes, including oat phytochrome A and Arabidopsis phytochrome B. |
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Keywords: | Cph1 cyanobacterial phytochrome 1 FR far-red light λmax the wavelength of maximum absorption MTS mitochondrial targeting sequence PCB phycocyanobilin PΦB phytochromobilin phyA phytochrome A phyB phytochrome B PIF3 phytochrome-interacting factor 3 Pr red-absorbing form of phytochrome Pfr far-red-absorbing form of phytochrome R red light |
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