Which one of the following statements concerning EcoRV, a restriction enzyme is

Question

Which one of the following statements concerning EcoRV, a restriction enzyme is incorrect? g. A Mg2+ ion activates water as part of the attack of the scissile bond. h. The cleavage occurs at sites of inverted repeats. i. EcoRV acts as a dimer demonstrating symmetry for the two-fold rotational symmetry of the DNA cognate. j. Mg2+ is held in place by covalently binding Asp residues. k. Non-cognate DNA is not cleaved because it is distorted by EcoRV. l. Hydrogen binding stabilizes the cognate DNA in the active site.

Explanation / Answer

The restriction endonuclease EcoRV can rapidly locate a short recognition site within long non-cognate DNA using ‘facilitated diffusion’. This process has long been attributed to a sliding mechanism, in which the enzyme first binds to the DNA via nonspecific interaction and then moves along the DNA by 1D diffusion. Recent studies, however, provided evidence that 3D translocations (hopping/jumping) also help EcoRV to locate its target site. Here we report the first direct observation of sliding and jumping of individual EcoRV molecules along nonspecific DNA. Using fluorescence microscopy, we could distinguish between a slow 1D diffusion of the enzyme and a fast translocation mechanism that was demonstrated to stem from 3D jumps. Salt effects on both sliding and jumping were investigated, and we developed numerical simulations to account for both the jump frequency and the jump length distribution. i. EcoRV acts as a dimer demonstrating symmetry for the two-fold rotational symmetry of the DNA cognate.

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