STABILIZATION OF THE NEUTRAL PROTEASE OF BACILLUS-STEAROTHERMOPHILUS BY REMOVAL OF A BURIED WATER MOLECULE

Using site-directed mutagenesis, Ala166 in the neutral protease of Bacillus stearothermophilus was changed into Ser. Model building and molecular dynamics simulations of the mutant enzyme indicated that the Ser hydroxyl group fits well in a cavity which contains a water molecule in the wild-type enzyme. The Ala166 --> Ser mutation was expected to exert a stabilizing effect because of the gain in entropy resulting from the release of a water molecular from the folded protein to the solvent. In addition, the hydrogen-bonding network around residue 166 was improved upon the mutation. As a result of this mutation the thermostability of the neutral protease was increased by 1.2 +/- 0.1-degrees-C.