Engineering proteins with tunable thermodynamic and kinetic stabilities.

摘要: It is widely recognized that enhancement of protein stability an important biotechnological goal. However, some applications at least, could actually benefit from being strongly dependent on a suitable environment variable, in such way enhanced or decreased be realized as required. In therapeutic applications, for instance, long shelf-life under storage conditions may convenient, but sufficiently fast degradation the after it has performed planned molecular task vivo avoid side effects and toxicity. Undesirable associated to high are also likely occur food-industry applications. Clearly, one fundamental factor involved here kinetic protein, which relates time-scale irreversible denaturation processes determined significant extent by free-energy barrier unfolding (the “separates” native state highly-susceptible-to-irreversible-alterations nonnative states). With appropriate experimental model, we show strong environment-dependencies thermodynamic stabilities can achieved using robust engineering. We use sequence-alignment analysis simple computational electrostatics design stabilizing destabilizing mutations, latter introducing interactions between like charges screened out salt. Our procedures lead naturally mutating regions mostly unstructured transition unfolding. As result, large salt effect our consensus plus charge-reversal variant translates into dramatic changes barrier: order years days low Certainly, concentration not expected biological systems vivo. Hence, proteins with salt-dependencies more those cases high-stability required only conditions. A plausible scenario inclusion liquid formulations will contribute shelf-life, while lower application help prevent potentially arise From general viewpoint, this work shows engineering electrostatic readily combined clarifies relevant aspects relation proteins. Proteins 2008. © 2007 Wiley-Liss, Inc.