Recently, the team of Professor Xu Yan and Yu Xiaowei from the Brewing Microbiology and Applied Enzymology Laboratory of Jiangnan University’s Bioengineering College has made important progress in the research of lipase propeptide. The cover of substrate selectivityby computational design” was published in the Journal of Agricultural and Food Chemistry (IF=4.192) (https://doi.org/10.1021/acs.jafc.1c0072)
lipase (EC 22.214.171.124) has good hydrolysis and synthesis activity on ester bonds, and is widely used in food, pharmaceutical, detergent and bio-energy related industries. Among them, fungal lipase has obvious advantages due to its high stability and wide substrate specificity. There is a propeptide domain in the structure of most fungal lipases, which is very important for the secretion, catalytic efficiency and substrate selectivity of the lipase. In this study, Rhizopus chinensis lipase (Rhizopus chinensislipase, RCL) was used as the research model, using metadynamics (metadynamics) simulation, classical molecular dynamics simulation, and site-directed mutagenesis to discuss in depth The peptide domain affects its function and the mechanism of action of substrate selectivity. The calculation results show that the propeptide domain significantly inhibits the movement of the RCL lid. Through dynamic cross-correlation analysis and independent gradient model analysis, a key region of inhibition (Val5-Thr10) is found on the leader peptide, and in this fragment Carry out rational design and determination of enzymatic reaction kinetics. The catalytic efficiency of the optimal lipase mutant is 7 times higher than that of the wild type, and it also shows the ability of the propeptide domain to change the chain length selectivity of the lipase substrate. After substrate expansion, it is found that this regulatory ability is also reflected in other substrates of triglycerides. In this study, computer simulation and site-directed mutagenesis are combined with each other to reveal the important role of the key region of the propeptide in improving the catalytic efficiency of lipase and the controllable change of substrate specificity. It is also an important factor for industrial enzymes that also have similar domains of the leader peptide. The design and transformation provide a theoretical basis.
Figure 1. Independent gradient model analysis of Rhizopus sinensis lipase propeptide and lid in pNPb(A) and pNPp( B) Interaction in the substrate system
Professor Yu Xiaowei is the corresponding author of this paper, and doctoral student Wang Shang is the first author of this paper. The above-mentioned research work was funded by the National Natural Science Foundation of China (32072162) and the National Double First-Class Discipline Construction Project of Light Industry Technology and Engineering (LITE2018-09).