Raquet X, Vanhove M, Lamotte-Brasseur J, Goussard S, Courvalin P, Frère J M
Centre d'Ingénierie des Protéines, Université de Liège, Sart-Tilman, Belgium.
Proteins. 1995 Sep;23(1):63-72. doi: 10.1002/prot.340230108.
The stability properties of six natural mutants of the TEM-1 beta-lactamase have been studied. The glutamate to lysine substitution at positions 104 and 240 stabilize the enzyme. Conversely, the G238S mutant's decreased stability might reflect an altered conformation of the active site and thus be related to the modified substrate profile. The relative stability of the R164S and R164H mutants is explained by the formation of a hydrogen bond between these residues and Asp-179 conferring a somewhat different structure to the omega loop and thus also explaining the extended substrate profile of these mutants. The loss of stability of the R164H mutant with increasing pH values can be explained by the titration of a hydrogen bond between the N delta of His-164 and the O delta of Asp-179. The properties of the G238S + E104K double mutant which is the most active against third-generation cephalosporins result from a balance of destabilizing and stabilizing substitutions, and their effects seem to be additive. The behavior of the R164S + E240K mutant might be explained on the basis of a similar compensation phenomenon.
已对TEM-1β-内酰胺酶的六个天然突变体的稳定性特性进行了研究。104位和240位的谷氨酸被赖氨酸取代使该酶更稳定。相反，G238S突变体稳定性的降低可能反映了活性位点构象的改变，因此与修饰后的底物谱有关。R164S和R164H突变体的相对稳定性可以通过这些残基与Asp-179之间形成氢键来解释，这赋予了ω环 somewhat different结构，因此也解释了这些突变体扩展的底物谱。R164H突变体随着pH值升高稳定性的丧失可以通过His-164的Nδ与Asp-179的Oδ之间氢键的滴定来解释。对第三代头孢菌素活性最强的G238S + E104K双突变体的特性是由不稳定和稳定取代的平衡导致的，它们的作用似乎是相加的。R164S + E240K突变体的行为可能基于类似的补偿现象来解释。
