Facilitated diffusion of the EcoRI DNA methyltransferase is described by a novel mechanism.
The contribution of nonspecific DNA to binding parameters (K(d), k(off), and k(on)) was determined for the EcoRI DNA methyltransferase under noncatalytic conditions. An increase in DNA size from 14 to 775 base pairs causes a 20-fold decrease in K(d), while k(off) remains constant over the same range. The calculated k(on) increases with longer substrates, consistent with a facilitated diffusion mechanism. However, the combined results deviate from the model developed to describe facilitated diffusion [Berg, O. G., Winter, R. B., & von Hippel, P. H. (1981) Biochemistry 20, 6929-6948]. Our results were successfully simulated using numerical integration of a kinetic scheme invoking protein dissociation via the ends of DNA. Consistent with this scheme, the methyltransferase dissociates more slowly from a circularized DNA molecule than from the identical linearized form. The simulation strategy correctly models our data with the methyltransferase and should be generally useful for routine modeling of facilitated diffusion involving protein-DNA systems.
Surby
M A
MA
Department of Chemistry, University of California, Santa Barbara 93106, USA.
Reich
N O
NO
D001483:Base Sequence; D002384:Catalysis; D004247:DNA / Q000737:chemistry* / Q000378:metabolism; D015246:Deoxyribonuclease EcoRI / Q000378:metabolism*; D004058:Diffusion; D008969:Molecular Sequence Data; D013379:Substrate Specificity; D013816:Thermodynamics
D004247:DNA; D015246:Deoxyribonuclease EcoRI