In vitro formation of a photoreversible adduct of phycocyanobilin and tobacco apophytochrome B.
The light-stable tobacco phytochrome apoprotein (PHYB) expressed in yeast can be assembled with phycocyanobilin to give a photoreversible adduct. The spectral properties of the reconstituted PHYB-phycocyanobilin species were determined by absorbacen and difference absorbance spectroscopies. The holoprotein exhibits absorbance maxima at 408 nm and 712 nm for the far-red-light-absorbing (Pfr) form and 356 nm and 658 nm for the red-light-absorbing (Pr) form. The ligation of the chromophores to the dimeric PHYB apoprotein resulted in a PHYB-phycocyanobilin adduct with the spectral properties of the Pr form. Kinetic analyses of the in vitro reconstitution for PHYB apoprotein under saturating concentrations of phycocyanobilin revealed a pseudo first-order rate constant of 2.8 x 10(-2)s.-1. The similarity with the reported rate constant for the reconstitution of light-labile phytochrome (PHYA) from oat [Li, L. & Lagarias, J.C. (1992) Phytochrome assembly, J. Biol. Chem. 267, 19,204-19,210] suggests that the mechanisms of chromophore attachment are probably very similar for PHYA and PHYB.
Kunkel
T
T
Institut für Biologie II, Universität Freiburg, Germany.
Tomizawa
K
K
Kern
R
R
Furuya
M
M
Chua
N H
NH
Schäfer
E
E
D001059:Apoproteins / Q000737:chemistry* / Q000235:genetics; D002478:Cells, Cultured; D003001:Cloning, Molecular; D004591:Electrophoresis, Polyacrylamide Gel; D007700:Kinetics; D052980:Phycobilins; D010798:Phycocyanin / Q000737:chemistry*; D010834:Phytochrome / Q000737:chemistry* / Q000235:genetics; D010947:Plants, Toxic; D011758:Pyrroles / Q000737:chemistry*; D012441:Saccharomyces cerevisiae; D013057:Spectrum Analysis; D045725:Tetrapyrroles; D014026:Nicotiana / Q000201:enzymology*
D001059:Apoproteins; D052980:Phycobilins; D011758:Pyrroles; D045725:Tetrapyrroles; C071907:apophytochrome; D010798:Phycocyanin; D010834:Phytochrome; C011069:phycocyanobilin