{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,3,19]],"date-time":"2026-03-19T17:34:33Z","timestamp":1773941673971,"version":"3.50.1"},"reference-count":37,"publisher":"Wiley","issue":"4","license":[{"start":{"date-parts":[[2006,12,12]],"date-time":"2006-12-12T00:00:00Z","timestamp":1165881600000},"content-version":"vor","delay-in-days":0,"URL":"http:\/\/onlinelibrary.wiley.com\/termsAndConditions#vor"}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":["Biotech &amp; Bioengineering"],"published-print":{"date-parts":[[2007,7]]},"abstract":"<jats:title>Abstract<\/jats:title><jats:p>Engineered extremely thermostable variants of the thermolysin\u2010like protease from <jats:italic>Bacillus stearothermophilus<\/jats:italic> possessing an introduced disulfide bond G8C\/N60C (double mutant, DM) and six additional amino acid substitutions in the exposed loop region 56\u201369 (Boilysin, BLN) have been probed with respect to stability toward water\u2010miscible organic solvents and detergents. The solvent concentrations where 50% of enzyme activity were irreversibly lost (C<jats:sub>50<\/jats:sub>) decreased in the order methanol\u2009&gt;\u2009 2\u2010propanol\u2009&gt;\u2009dimethylsulfoxide\u2009&gt;\u2009dioxane\u2009&gt;\u2009acetonitrile\u2009&gt;\u2009dimethylformamide\u2009&gt;\u2009acetone. The C<jats:sub>50<\/jats:sub> values were remarkably higher for the thermostable variants than for the wild\u2010type enzymes. Therefore, the stabilization of this loop region also protects the molecule from irreversible inactivation by solvents, and inactivation seems to follow principally the same mechanism as thermal inactivation. However, in contrast to thermal inactivation where the corresponding T<jats:sub>50<\/jats:sub> values of DM and BLN differed by 10 K, the differences of the C<jats:sub>50<\/jats:sub> values of DM and BLN were not significant. Detergents had great effects on proteolytic activities which were dependent on the individual detergent and its concentration, but mostly without significant differences between the enzyme variants. These effects were inactivating (SDS, sulfobetaine) or strongly activating (CTAB, CHAPS). Triton X\u2010100 and Tween 20 were activating or inactivating at low and high concentrations, respectively. In all detergents, stabilities of the enzymes were strongly decreased. However, the more thermostable variants were affected by the detergents to the same extent as the wild\u2010type enzymes suggesting that the mechanism of detergent inactivation is different from that of thermal inactivation. Biotechnol. Bioeng. 2007;97: 672\u2013679. \u00a9 2006 Wiley Periodicals, Inc.<\/jats:p>","DOI":"10.1002\/bit.21292","type":"journal-article","created":{"date-parts":[[2006,12,12]],"date-time":"2006-12-12T20:47:06Z","timestamp":1165956426000},"page":"672-679","source":"Crossref","is-referenced-by-count":32,"title":["The stability of engineered thermostable neutral proteases from <i>Bacillus stearothermophilus<\/i> in organic solvents and 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