THERMOPHILIC PROTEINS - STABILITY AND FUNCTION IN AQUEOUS AND ORGANIC-SOLVENTS

The molecular stability of thermophilic and hyperthermophilic enzymes generally reflects the growth temperatures of the parent organisms. Ex tracellular enzymes from the hyperthermophilic Archaea typically show very high levels of thermal stability and a number of enzymes with T-m values of greater than 100 degrees C have been reported. The mechanis ms responsible for high molecular stability are typically intrinsic ch aracteristics of the protein, as shown by the comparative stabilities of many native and recombinant proteins. However, some extrinsic stabi lisation mechanisms have been demonstrated. High levels of thermal sta bility are positively correlated with stability in the presence of oth er denaturing agents, including detergents and organic solvents. This correlation suggests a common denaturation pathway where molecular mob ility/flexibility is the prime determinant of susceptibility to irreve rsible denaturation. In single phase organic-aqueous solvents, protein destabilisation occurs via solvent-induced alteration to the protein hydration shell. However, correlations between protein stability and s olvent hydrophobicity are unreliable. In two-phase organic aqueous sys tems, interfacial denaturation predominates and is a function of both interfacial tension and interfacial surface area. Intracellular enzyme s are protected from interfacial denaturation but are potentially susc eptible to direct organic solvent effects, possibly depending on the r ole of the cell wall and cell membrane in the partitioning of the orga nic solvent into the cell cytoplasm. Immobilisation of thermophilic en zymes provides a method for enhancing both the thermal and solvent sta bilities of thermophilic and mesophilic enzymes. Multi-point covalent immobilisation to glyoxal-agarose enhances thermal stability and limit s protein protein inactivation mechanisms. Miscible organic solvents h ave a profound influence on the specificities of enzyme reactions. The presence of high concentrations of miscible organic solvents may indu ce gross changes in substrate specificity and/or more subtle alteratio ns in chiral selectivity. Correlations between the variation in enanti oselectivity and both solvent hydrophobicity and solvent dielectric co nstant have been demonstrated although some recent studies implicate t he formation of specific solvent-enzyme complexes which directly affec t reaction kinetics. (C) 1997 Elsevier Science Inc.