Thermostable esterase estUT1 of bacteria Ureibacillus thermosphaericus: the influence of additional processed domain TrxA on enzyme properties

Abstract

A gene of thermostable esterase of bacteria Ureibacillus thermosphaericus was expressed in strain E. coli BL21(DE3) comprised in domain TrxA-containing genetic construct pET32b-estUT1, under the control T7-promoter. The specific activity and relative thermostability of thus produced recombinant enzyme increased from 54.2 to 65.8 % (an hour incubation at 70 °C). The additional domain TrxA was discovered not to affect noticeably the pH optimum of the enzyme activity and its substrate specificity. In the absence of domain TrxA, the stability of estUT1 increased considerably in the presence of various chemicals including ethanol and methanol. The maximal catalytic activity (kcat/KM) of esterase equal to 280.0 s–1·mM–1 was observed in the absence of domain TrxA. Thus, introduction of an additional processed domain TrxA allows the enzyme to be secreted in the dissolved form but, on the other hand, the target protein becomes less thermostable.