Remarkable Improvement of Methylglyoxal Synthase Thermostability by His–His Interaction

Abstract

Lately it has been proposed that interaction between two positively charged side chains can stabilize the folded state of proteins. To further explore this point, we studied the effect of histidine-histidine interactions on thermostability of methylglyoxal synthase from Thermus sp. GH5 (TMGS). The crystal structure of TMGS revealed that His23, Arg22, and Phe19 are in close distance and form a surface loop. Here, two modified enzymes were produced by site-directed mutagenesis (SDM); one of them, one histidine (TMGS-HH(O)), and another two histidines (TMGS-HHH(O)) were inserted between Arg22 and His23 (H(O)). In comparison with the wild type, TMGS-HH(O) thermostability increased remarkably, whereas TMGS-HHH(O) was very unstable. To explore the role of His23 in the observed phenomenon, the original His23 in TMGS-HHH(O) was replaced with Ala (TMGS-HHA). Our data showed that the half-life of TMGS-HHA decreased in relation to the wild type. However, its half-life increased in comparison with TMGS-HHH(O). These results demonstrated that histidine-histidine interactions at position 23 in TMGS-HH(O) probably have the main role in TMGS thermostability.

Identification and Structural Analysis of Amino Acid Substitutions that Increase the Stability and Activity of Aspergillus niger Glucose Oxidase

“…On the other hand, single mutations H172K and H220D both showed higher thermal sensitivity than the wild type ( Fig 3A ). A likely explanation is that the pair H172/H220 represents a His-His motif, which has been shown that increases protein stability [ 23 – 25 ]. The loss of the His-His interaction would be compensated by the introduction of the salt bridge.…”