Kinetic analysis of DNA binding by the c-Myb DNA-binding domain using surface plasmon resonance.
Kinetics of the interaction of the c-Myb DNA-binding domain (R2R3) with its target DNA have been analyzed by surface plasmon resonance measurements. The association and dissociation rate constants between the standard R2R3, the Cys130 mutant substituted with Ile, and the cognate DNA are 2.3x10(5) M(-1) s(-1) and 2.6x10(-3) s(-1) at pH 7.5 and 20 degrees C, respectively. Kinetic analyses of the binding of the standard R2R3 to the non-cognate DNAs and those of the R2R3 mutant proteins to the cognate DNA showed that the reduction of the binding affinity was mainly due to an increase in the dissociation rate.
Oda
M
M
Biomolecular Engineering Research Institute, Suita, Osaka, Japan.
Furukawa
K
K
Sarai
A
A
Nakamura
H
H
D000818:Animals; D001665:Binding Sites; D004247:DNA / Q000737:chemistry / Q000378:metabolism*; D004268:DNA-Binding Proteins / Q000737:chemistry / Q000378:metabolism*; D007700:Kinetics; D051379:Mice; D011485:Protein Binding; D011518:Proto-Oncogene Proteins / Q000737:chemistry / Q000378:metabolism*; D020598:Proto-Oncogene Proteins c-myb; D020349:Surface Plasmon Resonance; D015534:Trans-Activators / Q000737:chemistry / Q000378:metabolism*
D004268:DNA-Binding Proteins; D011518:Proto-Oncogene Proteins; D020598:Proto-Oncogene Proteins c-myb; D015534:Trans-Activators; D004247:DNA