The organophosphorus hydrolase (OPH) has been used to degrade organophosphorus chemicals, as one of the most frequently used decontamination methods. Under chemical and thermal denaturing conditions, the enzyme has been shown to unfold. To utilize this enzyme in various applications, the thermal stability is of importance. The engineering of de novo disulphide bridges has been explored as a means to increase the thermal stability of enzymes in the rational method of protein engineering. In this study, Disulphide by Design software, homology modelling and molecular dynamics simulations were used to select appropriate amino acid pairs for the introduction of disulphide bridge to improve protein thermostability. The thermostability of the wild-type and three selected mutant enzymes were evaluated by half-life, delta G inactivation (ΔGi) and structural studies (fluorescence and far-UV CD analysis). Data analysis showed that half-life of A204C/T234C and T128C/E153C mutants were increased up to 4 and 24 min, respectively; however, for the G74C/A78C mutant, the half-life was decreased up to 9 min. For the T128C/E124C mutant, both thermal stability and Catalytic efficiency (kcat) were also increased. The half-life and ΔGi results were correlated to the obtained information from structural studies by circular dichroism (CD) spectrometry and extrinsic fluorescence experiments; as rigidity increased in A204C/T2234C and T128C/E153C mutants, half-life and ΔGi also increased. For G74C/A78C mutant, these parameters decreased due to its higher flexibility. The results were submitted a strong evidence for the possibility to improve the thermostability of OPH enzyme by introducing a disulphide bridge after bioinformatics design, even though this design would not be always successful.

No abstract

No abstract

Although functionally graded biomaterials (FGBMs) have recently been employed in the dental implants, the role of mechanical complications in the FGBMs, i.e., isotropicity or orthotopicity, has not been well understood. This study, hence, was aimed at investigating the effects of a special type of FGBM implant in stress distribution of bone-implant interface using finite element method. Meanwhile, the effects of simplifications, such as presuming isotropic material model instead of an orthotropic one for a jaw bone, and applying a small deflection effect as a replacement for a large deflection effect in the stress-strain calculation were also examined. The results revealed that the FGBM implants can diminish the maximum stress in the implant-bone interface. In addition, although the amount of maximum strain in the bone and implant were low, considering a small deflection effect instead of a large deflection one showed to have a considerable influence in the stress and the displacement of the implant-bone system. The orthotropic bone also indicated a large amount of stress compared with the isotropic one which implies the importance of material models in simulating the stresses and displacements of the implant-bone system. These results have implications not only for understanding the stresses and displacements in the implant-bone interface, but also for providing a comprehensive information for the biomechanical experts to pay enough attention to the material models being employed in their numerical simulations.