期刊
JOURNAL OF AGRICULTURAL AND FOOD CHEMISTRY
卷 69, 期 14, 页码 4263-4275出版社
AMER CHEMICAL SOC
DOI: 10.1021/acs.jafc.1c00721
关键词
lipase; specificity; molecular dynamic simulation; computational design; metadynamics
资金
- National Natural Science Foundation of China (NSFC) [32072162]
- Postgraduate Research & Practice Innovation Program of Jiangsu Provence [KYCX17_1421]
- High-end Foreign Experts Recruitment Program [GDT20153200044]
- National First-Class Discipline Program of Light Industry Technology and Engineering [LITE2018-09]
- 111Project [111-2-06]
In this study, the mechanism of a propeptide in fungal lipases was explored using Rhizopus chinensis lipase as a research model. Molecular dynamics simulations showed that the propeptide inhibited the lid movement of the lipase and identified a key region on the propeptide. Subsequent mutation experiments confirmed the important role of this region in catalytic efficiency and substrate specificity of the lipase.
Most fungal lipases contain a propeptide, which is very important for their function and substrate selectivity. In the present study, Rhizopus chinensis lipase (RCL) was used as a research model to explore the mechanism of the propeptide of the lipase. Conventional molecular dynamics (MD) and metadynamics simulations were used to explore the mechanism by which the propeptide affects the activity of the lipase, which was subsequently verified by mutation experiments. MD simulations indicated that the propeptide had an inhibitory effect on the lid movement of RCL and found a key region (Val5-Thr10) on the propeptide. Subsequently, site-directed mutations were created in this region. The mutations enhanced the lipase catalytic efficiency to 700% and showed the potential for the propeptide to shift the substrate specificity of RCL. The specificity and activity of RCL mutants also had similar trends to wild-type RCL toward triglycerides with varying chain lengths. The mutual corroboration of simulation and sitedirected mutagenesis results revealed the vital role of the key propeptide region in the catalytic activity and substrate specificity of the lipase.
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