... the unfolding.Calcium ions are the main contributory factor to enzyme stability The binding of calcium ions seems to be the mainsource of stabilization of the neutral protease fromB. stearothermophilus. ... (Stratagene, Heidelberg, Germany), the primers5¢-GTTTTGCCCGGCAGCTTGTTTACCGATGGCGACAACCAA-3¢ (forward) and 5¢-TTGGTTGTCGCCATCGGTAAACAAGCTGCCGGGCAAAAC-3¢ (reverse), and the wild-type gene of the ... assuming that inactivation is determinedby autoproteolysis as a consequence of rate-limitingunfolding. A separate kinetic analysis of conformation-al unfolding, however, is hampered by autoproteolysis.Recently,...