Biao Feng

Enhancing the thermal stability of inulin fructotransferase with high hydrostatic pressure

The thermal stability of inulin fructotransferase (IFTase) subjected to high hydrostatic pressure (HHP) was studied. The value of inactivation rate of IFTase in the range of 70-80°C decreased under the pressure of 100 or 200 MPa, indicating that the thermostability of IFTase under high temperature was enhanced by HHP. Far-UV CD and fluorescence spectra showed that HHP impeded the unfolding of the conformation of IFTase under high temperature, reflecting the antagonistic effect between temperature and pressure on IFTase. The new intramolecular disulfide bonds in IFTase were formed under a combination of HHP and high temperature. These bonds might be related to the stabilization of IFTase at high temperature. All the above results suggested that HHP had the protective effect on IFTase against high temperature.

Improving the catalytic behavior of inulin fructotransferase under high hydrostatic pressure

BACKGROUND The demand for difructose anhydride III (DFA III), a novel functional sweetener, is growing continuously. It is produced from inulin by inulin fructotransferase (IFTase). In this study, high hydrostatic pressure (HHP), as a clean technology, was first applied to further improve the catalytic efficiency of IFTase in the process. RESULTS The maximum activity of IFTase was obtained under 200 MPa at 60 °C. Meanwhile, HHP lowered the energy barrier necessary for the enzymatic reaction and decreased the volume between the reactants and the transition state. Under this condition, the optimal pH for the enzymatic reaction shifted from 5.5 to 6.0. The activity was further enhanced by 65.2% in the presence of 1.5 mol L(-1) NaCl. CONCLUSION The catalytic reaction of IFTase was performed under HHP for the first time. HHP, as a promising green technology for bioconversion, significantly accelerated the enzymatic reaction under the appropriate operational conditions.

Elucidation of pressure-induced lid movement and catalysis behavior of Rhizopus chinensis lipase

The changes of lid movement and catalysis behavior of Rhizopus chinensis lipase under high hydrostatic pressure treatment was studied. Molecular dynamics simulation showed that the lipase lid under pressure was partially opened at below 200MPa but got more closed at over 400MPa. The interfacial activation changed little at pressure below 400MPa but became marginal with the pressure increased to 500MPa. The lipase hydrolysis ability by high pressure treatment underwent a course of initial increasing then reducing with maximum activity obtained at 200MPa and 40°C. At moderate given pressure, the pressure treatment lowered the volume between the reactants and the transition state. Km decreased from 0.592 to 0.441mmol/L with pressure increasing from 0.1 to 200MPa. Meanwhile, vmax and Kcat increased 24.2% and 20% respectively. The change trend of reaction kinetic parameters under 400-500MPa was contrary to that under 0.1-200MPa.

Coupled effects of salt and pressure on catalytic ability of Rhizopus chinensis lipase

BACKGROUND Both high pressure and environmental factors could influence the catalytic abilities of enzymes. This work investigated coupled effects of pressure and salts on Rhizopus chinensis lipase (RCL) to provide significant information for its further applications. RESULTS The maximum activity of RCL was observed under 200 MPa at 40 °C. The highest activity was achieved at concentrations of 0.06-0.1 mol L-1 for tested salts. The effect of monovalent cations on RCL activity followed the Hofmeister series (K+ > Na+ > Li+ ) at 0.1 MPa but the order of Na+ and K+ was changed under 200 MPa. Meanwhile, the effects of anions did not follow the Hofmeister series. KCl slightly improved the thermostability of RCL at moderate concentration. At 60 °C, LiCl only stabilised RCL at 0.1 mol L-1 . The pre-transition unfolding point was shifted from 4.5 to 3.5 mol L-1 with pressure increasing from 0.1 to 600 MPa. In addition, KCl could not change the lipases extrinsic fluorescence evolution versus pressure. CONCLUSION Pressure and salts could improve catalytic ability and stability of RCL under appropriate conditions. The effect of high pressure on RCL was influenced by salts. Meanwhile salts cannot prevent high pressure-induced damage to RCL.