Fluorescence emission and polarization for analyzing binding of ruthenium metalloglycocluster to lectin and tetanus toxin C-fragment.
We have designed and synthesized ruthenium complexes bearing clustered galactose Ru(bpy-2Gal)(3) and glucose Ru(bpy-2Glc)(3). Changes in fluorescence emission (FE) and fluorescence polarization (FP) of the metalloglycoclusters were measured by adding each lectin (peanut agglutinin (PNA), Ricinus communis agglutinin 120 (RCA), concanavalin A (ConA), or wheat germ agglutinin (WGA)) or tetanus toxin c-fragment (TCF). Following the addition of PNA and ConA, the FE spectra of Ru(bpy-2Gal)(3) and Ru(bpy-2Glc)(3) showed new emission peaks, respectively. In addition, Ru(bpy-2Gal)(3) and Ru(bpy-2Glc)(3) exclusively increased the FP values by addition of PNA and ConA. Since other combinations of the metalloglycoclusters and lectin caused little change, specific bindings of galactose to PNA and glucose to ConA were confirmed by the FE and FP measurement. From the FP analyses, the dissociation constants (K(d)) of Ru(bpy-2Gal)(3) to PNA and Ru(bpy-2Glc)(3) to ConA were calculated to be ca. 6.1 x 10(-6) M and 1.8 x 10(-5) M. Furthermore, the FP analyses proved specific binding of Ru(bpy-2Gal)(3) to TCF.
Okada
Tomoko
T
Graduate School of Bionics, Tokyo University of Technology, Katakura, Hachioji, Tokyo 192-0982, Japan. tomoko@bs.teu.ac.jp.
Makino
Taro
T
Minoura
Norihiko
N
D001665:Binding Sites; D003017:Clostridium tetani / Q000737:chemistry; D003208:Concanavalin A / Q000737:chemistry / Q000378:metabolism; D005690:Galactose / Q000138:chemical synthesis / Q000737:chemistry* / Q000378:metabolism; D005947:Glucose / Q000138:chemical synthesis / Q000737:chemistry* / Q000378:metabolism; D008958:Models, Molecular; D015394:Molecular Structure; D019887:Peanut Agglutinin / Q000737:chemistry / Q000378:metabolism; D010446:Peptide Fragments / Q000737:chemistry / Q000378:metabolism*; D037121:Plant Lectins / Q000737:chemistry / Q000378:metabolism*; D010944:Plants / Q000737:chemistry; D011485:Protein Binding; D017975:Ruthenium Compounds / Q000138:chemical synthesis / Q000737:chemistry* / Q000378:metabolism*; D013050:Spectrometry, Fluorescence; D013744:Tetanus Toxin / Q000737:chemistry / Q000378:metabolism*
D019887:Peanut Agglutinin; D010446:Peptide Fragments; D037121:Plant Lectins; C017707:Ricinus communis agglutinin-1; D017975:Ruthenium Compounds; D013744:Tetanus Toxin; C040635:tetanus toxin fragment C; D003208:Concanavalin A; D005947:Glucose; D005690:Galactose