Effect of high hydrostatic pressures on 20S proteasome activity

The 20S proteasome is the catalytic core of the ubiquitin proteolytic pathw ay, which is implicated in many cellular processes. The cylindrical structu re of this complex consists of four stacked rings of seven subunits each. T he central cavity is formed by two beta catalytic subunit rings in which pr otein substrates are progressively degraded. The 20S proteasome is isolated in a latent form which can be activated in vitro by various chemical and p hysical treatments. In this study, the effects of high hydrostatic pressure s on 20S proteasome enzymatic activity were investigated. When proteasomes were subjected to increasing hydrostatic pressures, a progressive loss of p eptidase activities was observed between 75 and 150 MPa. The inactivation a lso occurred when proteasomes were pressurized in the presence of synthetic peptide substrates; this may be the result of the dissociation of the 20S particle into its subunits under pressure, as was shown by PAGE. Pressurize d proteasomes also lost their caseinolytic activity. In contrast, in the pr esence of casein, the pressure-induced inactivation and the dissociation of the 20S particles were prevented. In addition, in comparison to that obser ved at atmospheric pressure, their caseinolytic activity was increased unde r pressure. Following depressurization, the caseinolytic activity returned to basal levels but was further enhanced following an additional pressuriza tion treatment. Thus, the structure of the 20S pal-tide exhibits a certain degree of plasticity. This pressure-induced activation of the 20S proteasom e is discussed in relation to its hollow structure, its currently accepted proteolytic mechanism and the general effect of high pressures on the bioch emical reactions and structures of biopolymers.