The methylotrophic yeast Pichia pastoris synthesizes a functionally active chromophore precursor of the plant photoreceptor phytochrome.
Induction of the expression of an algal phytochrome cDNA in the methylotrophic yeast Pichia pastoris led to time-dependent formation of photoactive holophytochrome without the addition of exogenous bilins. Both in vivo and in vitro difference spectra of this phytochromic species are very similar to those of higher plant phytochrome A, supporting the conclusion that this species possesses a phytochromobilin prosthetic group. Zinc blot analyses confirm that a bilin chromophore is covalently bound to the algal phytochrome apoprotein. The hypothesis that P. pastoris contains phytochromobilin synthase, the enzyme that converts biliverdin IX alpha to phytochromobilin, was also addressed in this study. Soluble extracts from P. pastoris were able to convert biliverdin to a bilin pigment, which produced a native difference spectrum upon assembly with oat apophytochrome A. HPLC analyses confirm that biliverdin is converted to both 3E- and 3Z-isomers of phytochromobilin. These investigations demonstrate that the ability to synthesize phytochromobilin is not restricted to photosynthetic organisms and support the hypothesis of a more widespread distribution of the phytochrome photoreceptor.
Wu
S H
SH
Section of Molecular and Cellular Biology, University of California, Davis 95616, USA.
Lagarias
J C
JC
D001664:Biliverdine / Q000031:analogs & derivatives* / Q000096:biosynthesis; D010088:Oxidoreductases / Q000302:isolation & purification*; D010834:Phytochrome / Q000096:biosynthesis* / Q000235:genetics; D010843:Pichia / Q000378:metabolism*; D011994:Recombinant Proteins / Q000096:biosynthesis; D015032:Zinc / Q000032:analysis
D011994:Recombinant Proteins; D010834:Phytochrome; C075811:phytochromobilin; D010088:Oxidoreductases; C071703:phytochromobilin synthase; D015032:Zinc; D001664:Biliverdine