4.8 Article

Molecular Insights into the Enhanced Performance of EKylated PETase Toward PET Degradation

Journal

ACS CATALYSIS
Volume 11, Issue 12, Pages 7358-7370

Publisher

AMER CHEMICAL SOC
DOI: 10.1021/acscatal.1c01062

Keywords

polyethylene terephthalate (PET); PETase; zwitterionic polypeptide fusion; PET degradation; molecular mechanism

Funding

  1. National Key Research and Development Program of China [2018YFA0900702]
  2. National Natural Science Foundation of China [21621004]

Ask authors/readers for more resources

The fusion of a zwitterionic polypeptide to PETase has been shown to enhance catalytic performance and increase product release in the degradation of highly crystallized PET films. Structural analysis and molecular simulations reveal the enhanced stability and substrate binding properties of EKylation-modified PETases.
The accumulation of polyethylene terephthalate (PET) in the environment has brought an enormous threat to the global ecosystem. Although the recently reported PET hydrolase (PETase) displays an efficient decomposition to PET, the low activity and thermostability limit its practical applications. Herein, we introduce a biomodification strategy by fusing a zwitterionic polypeptide (5-30 kDa) consisting of alternating-charged glutamic acid (E) and lysine (K) residues to the C-terminus of PETase and find that increasing the fusion peptide length leads to the improved catalytic performance. The product release in the degradation of highly crystallized PET films (45.2% in crystallinity) by PETase-EK30 is promoted by over 11 times as compared with PETase. The molecular mechanism of the enhanced catalytic performance is investigated via structural analysis, substrate binding, and molecular simulations. Characterizations of the secondary and tertiary structures verify a strengthened structural stability of the EKylated PETases. Synchronous fluorescence spectra indicate a more open substrate-binding pocket after EKylation. MD simulations of enzyme-substrate complexes support that the EKylation induces the exposure of hydrophobic amino acids (W185, I208, and W159) in the substrate-binding pocket and the rotation of the benzene ring of Y87, which promote the substrate binding kinetics. This leads to the enhanced substrate affinity, exactly represented by the increased association constant and the decreased binding free energy. Besides, a shortened catalytic distance is observed from the MD simulations, which might also contribute to the enhanced catalytic activity toward PET degradation. The molecular insights into the enhanced enzyme performance would benefit in extending the application of the EKylation strategy in various enzymes.

Authors

I am an author on this paper
Click your name to claim this paper and add it to your profile.

Reviews

Primary Rating

4.8
Not enough ratings

Secondary Ratings

Novelty
-
Significance
-
Scientific rigor
-
Rate this paper

Recommended

No Data Available
No Data Available