PRESSURE-INDUCED INACTIVATION OF ESCHERICHIA-COLI BETA-GALACTOSIDASE - INFLUENCE OF PH AND TEMPERATURE

In order to assess the feasibility of a high-pressure immunodesorption process using a beta-galactosidase-anti-beta-galactosidase complex as a model, the influence of high hydrostatic pressure on the inactivati on of E. coli beta-galactosidase has been investigated. The irreversib le activity loss of beta-galactosidase was studied as a function of pH and temperature for pressures comprised between atmospheric pressure and 500 megapascal (MPa; 1 MPa = 10 bar). This enabled us to establish a practical pressure-temperature diagram of stability for this enzyme . The stability domains determined thus appeared to be strongly depend ent on the pH under atmospheric pressure of the phosphate buffer emplo yed for pressurisation. Therefore, to interpret meaningfully this resu lt, the influence of pressure on the pH-activity curve of beta-galacto sidase was investigated by using a high-pressure stopped-flow device. It appeared that the pH-activity curve of this enzyme was also reversi bly affected by pressures lower than 150 MPa. An interpretation of the se results in relation to the high-pressure induced changes of ionisat ion constants is proposed. For our practical purpose, the implications for the elaboration of a high-pressure immunodesorption process using beta-galactosidase as a tag, are discussed.