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The α-glycosidase from Xanthomonas campestris (XgtA) can specifically catalyze the transglycosylation reactions, thus can be applied for the enzymatic synthesis of α-glycosides. However, the low thermal stability of XgtA has been a bottleneck for its industrial application. In this research, a combined semi-rational design strategy was used based on analysis of the α-helices and β-turns of XgtA, FireProt prediction of thermostable mutants and molecular dynamic simulations for improving the thermostability. The key positions with a significant impact on the thermal stability of XgtA were identified and the effect of proline substitution was tested. Five single-point mutants V167P, A177P, A210P, A220P, T345P, and a combined mutant Mut5 were obtained with improved thermal stability. Mut5 displayed a 3.06-fold increase in time of half-life at 45°C without impairing its initial hydrolytic activity. Molecular dynamics simulations revealed that changes in the flexibility of amino acid residues, newly-forming hydrogen bonding networks and hydrophobic interactions were responsible for the improved thermostability. These results suggest that proline substitution of key flexible positions is an effective strategy for improving enzyme thermostability. [ABSTRACT FROM AUTHOR]

The adhesion force between metal-based antibacterial coatings and polymer substrate decides the effectiveness and durability of antibacterial performance. In this study, the silicone rubber substrate was modified by molecular bridge, i.e., polyvinylpyrrolidone, chitosan and (3-mercaptopropyl) trimethoxysilane, to provide 'anchors' which captured Ag during electroless plating, and Ag coatings were resultantly produced. Different molecular bridge offered different adhesion. The spread plate counting, fluorescent staining assay and bacterial growth kinetics test showed that the antibacterial and anti-biofilm performance against Escherichia coli and Staphylococcus aureus was directly related to coating-substrate adhesion strength. A relative weaker adhesion resulted in rapid bactericidal and bacteria growth inhibition effect, and vice versa. The inductively coupled plasma atomic emission spectrometry indicated that relative weak adhesion provided fast Ag release, and strong adhesion gave lower Ag cumulative concentration, which may be attributed to different bonding strength. The coatings displayed sustained-release kinetics with the maximum cumulative Ag concentration of only 0.45 mg/L after 45 days of immersion in PBS solution, which was far below the human cell toxicity concentration. Nevertheless, all tested Ag coatings exhibited effective and long-lasting antibacterial properties. The tailored adhesion and consequent different antibacterial effect provided a choice for clinical applications. [ABSTRACT FROM AUTHOR]