Rational surface charge engineering of haloalkane dehalogenase for boosting the enzymatic performance in organic solvent solutions

Biocatalysis in organic solvents(OSs)has numerous important applications,but native enzymes in OSs often exhibit limited catalytic performance.Herein,we proposed a computation-aided surface charge engineering strategy to improve the catalytic performance of haloalkane dehalogenase DhaA in OSs based on the energetic analysis of substrate binding to the DhaA surface.Several variants with enhanced OS resistance were obtained by replacing negative charged residues on the surface with positive charged residue(Arg).Particularly,a four-substitution variant E16R/E93R/E121R/E257R exhibited the best catalytic performance(five-fold improvement in OS resistance and seven-fold half-life increase in 40%(vol)dimethylsulfoxide).As a result,the overall catalytic performance of the variant could be at least 26 times higher than the wild-type DhaA.Fluorescence spectroscopy and molecular dynamics simulation studies revealed that the residue substitution mainly enhanced OS resistance from four aspects:(a)improved the overall structural stability,(b)increased the hydrophobicity of the local microenvironment around the catalytic triad,(c)enriched the hydrophobic substrate around the enzyme molecule,and(d)lowered the contact frequency between OS molecules and the catalytic triad.Our findings validate that computationaided surface charge engineering is an effective and ingenious rational strategy for tailoring enzyme performance in OSs.

Real-time Control Oriented HCCI Engine Cycle-to-cycle Dynamic Modelling

For homogeneous charge compression ignition （HCCI） combustion, the auto-ignition process is very sensitive to in-cylinder conditions, including in-cylinder temperature, in-cylinder components and concentrations. Therefore, accurate control is required for reliable and efficient HCCI combustion. This paper outlines a simplified gasoline-fueled HCCI engine model implemented in Simulink environment. The model is able to run in real-time and with fixed simulation steps with the aim of cycle-to-cycle control and hardware- in-the-loop simulation. With the aim of controlling the desired amount of the trapped exhaust gas recirculation （EGR） from the previous cycle, the phase of the intake and exhaust valves and the respective profiles are designed to vary in this model. The model is able to anticipate the auto-ignition timing and the in-cylinder pressure and temperature. The validation has been conducted using a comparison of the experimental results on Ricardo Hydro engine published in a research by Tianjin University and a JAGUAR V6 HCCI test engine at the University of Birmingham. The comparison shows the typical HCCI combustion and a fair agreement between the simulation and experimental results.