HEAT-INDUCED TRANSLOCATION OF CYTOPLASMIC BETA-GALACTOSIDASE ACROSS INNER MEMBRANE OF ESCHERICHIA-COLI

The behaviors of heat-induced translocation of cytoplasmic beta-galact osidase to periplasm across the inner membrane of Escherichia coli cel ls were investigated in order to apply such phenomena to the process f or production and separation of intracellular biomolecules. The heat s tress was found to induce translocation of cytoplasmic beta-galactosid ase (beta-gal) together with reduction of the amounts of intracellular soluble proteins and formation of their inactive aggregates. The tran slocation of beta-gal was then analyzed using (a) the location factor of beta-gal (LFG), which meant enzyme location in the cells and could be determined from the kinetic analysis of enzyme release process, and (b) the percentage of beta-gal activity in periplasm after solublizin g the outer membrane of E. coli cells by lysozyme/EDTA treatment. The LFG values were maximized when cells were stressed at the temperature of 42-47 degrees C. From the results on the surface properties of both beta-gal and cell membrane under the heat stress, it is suggested tha t (1) the conformational change of cytoplasmic oligomeric beta-gal to the partially dissociated and/or unfolded state with higher local hydr ophobicity, (2) the increase in membrane fluidity of inner membrane, ( 3) the enhancement of hydrophobic interaction between lipid and protei n, and (4) the inhibition of its translocation by GroEL restabilizing the proteins could underlie the heat-induced translocation of beta-gal across the inner membrane. The possibility to apply the heat-induced translocation of beta-gal for the enhancement of the target selectivit y at the process upstream is finally presented.