Purification and cDNA cloning of cytokinin-specific binding protein from mung bean (Vigna radiata).
Synthetic urea derivatives such as N-phenyl-N'-(4-pyridyl)urea (4PU) and N-(2-chloro-4-pyridyl)-N'-phenylurea (4PU30) have strong cytokinin activities. Using tritiated 4PU30 as a probe, we previously established the presence of a cytokinin-specific binding protein (CSBP) of high affinity (Ka for 4PU30 = 4x10(10) M(-1)) in the soluble fraction of etiolated mung bean seedlings [Nagata, R., Kawachi, E., Hashimoto, Y. & Shudo, K. (1993) Biochem. Biophys. Res. Commun. 191, 543-549]. In this report, we purified CSBP by the use of 4PU-Sepharose 4B, an affinity gel liganded with 4PU. We determined partial amino acid sequences of CSBP and isolated its cDNA by reverse-transcription (RT) PCR. The cDNA encoded a protein with a calculated molecular mass of 17 kDa. A data base homology search revealed that CSBP is a novel member of a major pollen allergen/pathogenesis-related protein family. Recombinant CSBP was expressed in Escherichia coli and was confirmed to bind specifically to cytokinins.
Fujimoto
Y
Y
Institute of Molecular and Cellular Biosciences, University of Tokyo, Japan.
Nagata
R
R
Fukasawa
H
H
Yano
K
K
Azuma
M
M
Iida
A
A
Sugimoto
S
S
Shudo
K
K
Hashimoto
Y
Y
D000595:Amino Acid Sequence; D029681:Arabidopsis Proteins; D001483:Base Sequence; D001667:Binding, Competitive; D002352:Carrier Proteins / Q000737:chemistry / Q000235:genetics*; D002846:Chromatography, Affinity / Q000379:methods; D003001:Cloning, Molecular; D003583:Cytokinins / Q000378:metabolism; D007887:Fabaceae / Q000737:chemistry*; D008969:Molecular Sequence Data; D010671:Phenylurea Compounds / Q000378:metabolism; D010802:Phylogeny; D010940:Plant Proteins; D010946:Plants, Medicinal; D011485:Protein Binding; D011725:Pyridines / Q000378:metabolism; D011994:Recombinant Proteins / Q000378:metabolism; D012685:Sepharose / Q000031:analogs & derivatives; D017422:Sequence Analysis, DNA; D017386:Sequence Homology, Amino Acid
D029681:Arabidopsis Proteins; D002352:Carrier Proteins; D003583:Cytokinins; D010671:Phenylurea Compounds; D010940:Plant Proteins; D011725:Pyridines; D011994:Recombinant Proteins; D012685:Sepharose; C116093:N-(2-chloro-4-pyridyl)-N'-phenylurea