title: Expression of recombinant full-length plant phytochromes assembled with phytochromobilin in Pichia pastoris.
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.
authors.affiliation: Department of Biotechnology and Kumho Life Science Laboratory, Chonnam National University, Gwangju 500-757, Republic of Korea.
authors.forename: Ah-Young
authors.initials: AY
authors.lastname: Shin
authors.affiliation: Department of Biotechnology and Kumho Life Science Laboratory, Chonnam National University, Gwangju 500-757, Republic of Korea.
authors.forename: Yun-Jeong
authors.initials: YJ
authors.lastname: Han
authors.affiliation: Faculty of Biotechnology and Subtropical Horticulture Research Institute, Jeju National University, Jeju 690-756, Republic of Korea.
authors.forename: Pill-Soon
authors.initials: PS
authors.lastname: Song
authors.affiliation: Department of Biotechnology and Kumho Life Science Laboratory, Chonnam National University, Gwangju 500-757, Republic of Korea. Electronic address: kimji@chonnam.ac.kr.
authors.forename: Jeong-Il
authors.initials: JI
authors.lastname: Kim
mesh_terms: D000595:Amino Acid Sequence; D001059:Apoproteins / Q000096:biosynthesis / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D001664:Biliverdine / Q000031:analogs & derivatives* / Q000378:metabolism; D015870:Gene Expression; D005818:Genetic Engineering / Q000379:methods*; D051547:Heme Oxygenase-1 / Q000235:genetics; D008928:Mitochondria / Q000378:metabolism; D008969:Molecular Sequence Data; D010834:Phytochrome / Q000096:biosynthesis / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism*; D010843:Pichia / Q000235:genetics*; D021381:Protein Transport; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism*
chemical_list: D001059:Apoproteins; D011994:Recombinant Proteins; D010834:Phytochrome; C075811:phytochromobilin; D051547:Heme Oxygenase-1; D001664:Biliverdine
keywords: Bilin reductase; Chromophore; Heme oxygenase; Holophytochrome; Phytochromobilin