Recombinant expression and inhibition mechanism analysis of pectin methylesterase from Aspergillus flavus
Xiuping Jiang1, Qiulei Jia, Lei Chen
1School of Life Science & Biotechnology, Dalian University of Technology, Dalian, China.
Abstract:
Phytopathogenic microorganisms can produce pectin methylesterase (PME) to degrade plant cell walls during plant invasion. This enzyme is thought to be a virulence factor of phytopathogens. In this work, PME from Aspergillus flavus (AFPME) was expressed in Pichia pastoris and an in vitro inhibitor study was performed. The purified AFPME with a yield of 52.2% was resolved as one band with a molecular mass of c. 40 kDa by SDS-PAGE. Optimal activity of the enzyme occurred at a temperature of 55 °C and a pH of 4.8. Epigallocatechin gallate (EGCG) strongly inhibited the activity of recombinant AFPME. The molecular docking analysis indicated that EGCG could form hydrogen bonds and π-π interactions with some amino acid residues in the active site of AFPME. Our studies provide a novel strategy for the control of the plant invasion of A. flavus.
Insights
This study purified Aspergillus flavus pectin methylesterase (PME) and found that epigallocatechin gallate (EGCG) strongly inhibits its activity. This discovery offers a new strategy for controlling fungal plant invasions.
Area of Science:
- Plant Pathology
- Enzymology
- Biochemistry
Background:
- Phytopathogenic microorganisms utilize pectin methylesterase (PME) to degrade plant cell walls, facilitating invasion.
- PME is considered a significant virulence factor in plant pathogens.
Purpose of the Study:
- To express and purify PME from Aspergillus flavus (AFPME).
- To investigate the inhibitory effects of epigallocatechin gallate (EGCG) on AFPME activity.
- To explore potential molecular interactions between EGCG and AFPME.
Main Methods:
- Recombinant expression of AFPME in Pichia pastoris.
- Enzyme purification and characterization (SDS-PAGE, optimal temperature and pH determination).
- In vitro enzyme inhibition assays and molecular docking analysis.
Main Results:
- Purified AFPME (approx. 40 kDa) showed optimal activity at 55°C and pH 4.8.
- Epigallocatechin gallate (EGCG) demonstrated strong inhibition of AFPME.
- Molecular docking revealed hydrogen bonds and π-π interactions between EGCG and AFPME active site residues.
Conclusions:
- Epigallocatechin gallate (EGCG) is a potent inhibitor of Aspergillus flavus pectin methylesterase (AFPME).
- The findings suggest a novel approach for managing plant invasions by A. flavus using EGCG.
