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An efficient peptide ligase engineered from a bamboo asparaginyl endopeptidase
Xin-Bo Wang1, Cong-Hui Zhang1, Teng Zhang1
1Research Center for Translational Medicine at East Hospital, School of Life Sciences and Technology, Tongji University, Shanghai, China.
The FEBS Journal
|March 25, 2024
Summary
Researchers discovered bamboo-derived asparaginyl endopeptidases (AEPs) with potent peptide ligase activity. An engineered mutant, [G238V]BmAEP1, offers enhanced control for protein labeling and cyclic peptide synthesis.
Area of Science:
- Biochemistry
- Molecular Biology
- Enzymology
Background:
- Asparaginyl endopeptidases (AEPs) from plants show promise as peptide ligases for protein labeling and cyclic peptide synthesis.
- A novel NanoLuc Binary Technology (NanoBiT)-based assay was developed to discover new AEP-type peptide ligases.
Purpose of the Study:
- To screen bamboo species for AEP-type peptide ligase activity.
- To identify and engineer a bamboo-derived AEP for improved protein ligation and peptide cyclization applications.
Main Methods:
- Screening of 61 bamboo species using a NanoBiT-based assay.
- Identification of BmAEP1 from Bambusa multiplex via transcriptomic sequencing.
- Site-directed mutagenesis and a novel protease-assisted activation method for engineered BmAEP1.
Main Results:
- AEP-type peptide ligase activity was detected in all 61 screened bamboo species.
- The identified BmAEP1 exhibited both high ligase and hydrolytic activity.
- Engineered [G238V]BmAEP1, activated via a protease-assisted method, showed high ligase activity with reduced hydrolysis, enabling efficient protein ligation and peptide cyclization.
Conclusions:
- Bamboo species are a rich source of AEP-type peptide ligases.
- The engineered [G238V]BmAEP1 provides a versatile and controllable tool for biotechnological applications.
- This study expands the toolkit for advanced protein engineering and synthetic chemistry.
