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AFM protein-protein interactions within the EcoR124I molecular motor

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Dynamic Force Spectroscopy (DFS), an Atomic Force Microscopy (AFM) technique, has been used to investigate the interaction between the HsdR subunit and the core methylase (MTase) of the Type I Restriction-Modification (R-M) enzyme EcoR124I. Such systems are of interest in bionanotechnology owing to their ability to translocate DNA, thus acting as molecular motors. Forces between a glutathione S-transferase (GST)-HsdR(PrrI) motor subunit attached to an AFM tip using a polyethylene gycol linker and the core MTase on poly-L-lysine pre-treated mica were measured at different loading rates. In the absence of an applied force, the position of energy barrier xdiss, bond dissociation rate kdiss(0) and lifetime of the bond (0) were calculated to be 1.35±0.17 nm, 0.16 s-1 and 6.3 s, respectively. The kdiss(0) value was a little lower than that obtained from magnetic tweezers (0.4 s-1), suggesting that the thermodynamic equilibrium may be affected by the presence of DNA. This work demonstrates that kinetic data concerning protein-protein interactions between subunits within Type I R-M enzymes are accessible via AFM. Such information is important for structure elucidation and the development of nanodevices.
Original languageEnglish
Pages (from-to)6358-6363
Number of pages6
JournalSoft Matter
Volume8
Issue number23
DOIs
Publication statusPublished - 2012

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