A fluorescence correlation spectroscopy study of ligand interaction with cytokinin-specific binding protein from mung bean.
Cytokinins are essential plant hormones that regulate numerous physiological processes. Recently, a protein was identified in mung bean (Vigna radiata) and characterized as a cytokinin-specific binding protein (VrCSBP). Fluorescence correlation spectroscopy was used to investigate the interaction between VrCSBP and its ligands. The synthetic cytokinin, N-phenyl-N'-(4-pyridyl) urea, was labeled with two fluorophores, 7-nitro-2,1,3-benzoxadiazole and rhodamine B. Protein-ligand binding was analyzed in an equilibrium saturation binding experiment and confirmed by the competition assay. Surprisingly, it was found that VrCSBP binds not only to cytokinins, but also to gibberellins. In addition, in the presence of natural cytokinins and gibberellins, two populations of VrCSBP that differ in their diffusion coefficients were detected. The diffusion coefficients of these two populations could be related to mono- and dimeric states, which suggests a new mode of operation in ligand binding by VrCSBP, in which dimerization induced by natural ligands enhances the ligand binding capacity of the protein.
Zawadzki
Paweł
P
Department of Molecular and Cellular Biology, Faculty of Biology, Adam Mickiewicz University, Umultowska 89, 61-614 Poznań, Poland. zawadzki@amu.edu.pl
Slósarek
Genowefa
G
Boryski
Jerzy
J
Wojtaszek
Przemysław
P
D009327:4-Chloro-7-nitrobenzofurazan / Q000737:chemistry; D002352:Carrier Proteins / Q000378:metabolism*; D003583:Cytokinins / Q000378:metabolism*; D007887:Fabaceae / Q000737:chemistry; D005456:Fluorescent Dyes / Q000737:chemistry; D005875:Gibberellins / Q000378:metabolism; D008024:Ligands; D010671:Phenylurea Compounds / Q000737:chemistry; D010940:Plant Proteins / Q000378:metabolism*; D011485:Protein Binding; D055503:Protein Multimerization; D011725:Pyridines / Q000737:chemistry; D012235:Rhodamines / Q000737:chemistry
D002352:Carrier Proteins; D003583:Cytokinins; D005456:Fluorescent Dyes; D005875:Gibberellins; D008024:Ligands; D010671:Phenylurea Compounds; D010940:Plant Proteins; D011725:Pyridines; D012235:Rhodamines; D009327:4-Chloro-7-nitrobenzofurazan; C029773:rhodamine B