Reversible Inhibition of EcoR I with Hydrostatic Pressure

The ability of EcoR I to cut at its recognition sequence in the plasmid pBR 322 was monitored as a function of hydrostatic pressure. EcoR I is completely inhibited at 200 MPa (atmospheric pressure = 0.1 MPa); however, the enzyme regains its original activity upon return to atmospheric pressure. At lower pressure the enzyme cuts normally but more slowly; there was no evidence of EcoR* activity at elevated pressures or upon return to atmospheric pressure. Assuming the reduction in activity is due exclusively to the decreased free energy of interaction between the enzyme and DNA the molar volume change (DELTAV) is estimated to be >= 85 cm sup 3 mol sup (-1). A positive DELTAV implies that the DNA-protein complex becomes energetically less favorable with increasing pressure because it occupies more volume than the individual species in solution. This volume change arises on account of differences in the extent of solvation of the bound and free forms; the pressures employed are much too low to cause any change in the intrinsic volume of the molecules.