Domain Swapping between AtACS7 and PpACL1 Results in Chimeric ACS-like Proteins with ACS or Cβ-S Lyase Single Enzymatic Activity
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Published:2023-02-03
Issue:3
Volume:24
Page:2956
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ISSN:1422-0067
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Container-title:International Journal of Molecular Sciences
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language:en
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Short-container-title:IJMS
Author:
Xu Chang1, Sun Lifang1, Mei Yuanyuan1, Sun Gongling1, Li Wenjing1, Wang Dan1, Li Xin2, Wang Ning Ning1ORCID
Affiliation:
1. College of Life Sciences, College of Agricultural Sciences, Tianjin Key Laboratory of Protein Sciences, Nankai University, Tianjin 300071, China 2. State Key Laboratory of Medicinal Chemical Biology, Frontiers Science Center for Cell Responses, College of Life Sciences, Nankai University, Tianjin 300071, China
Abstract
The gaseous hormone ethylene plays a pivotal role in plant growth and development. In seed plants, the key rate-limiting enzyme that controls ethylene biosynthesis is ACC synthase (ACS). ACS has, for a long time, been believed to be a single-activity enzyme until we recently discovered that it also possesses Cβ-S lyase (CSL) activity. This discovery raises fundamental questions regarding the biological significance of the dual enzymatic activities of ACS. To address these issues, it is highly necessary to obtain ACS mutants with either ACS or CSL single activity. Here, domain swapping between Arabidopsis AtACS7 and moss CSL PpACL1 were performed. Enzymatic activity assays of the constructed chimeras revealed that, R10, which was produced by replacing AtACS7 box 6 with that of PpACL1, lost ACS but retained CSL activity, whereas R12 generated by box 4 substitution lost CSL and only had ACS activity. The activities of both chimeric proteins were compared with previously obtained single-activity mutants including R6, AtACS7Q98A, and AtACS7D245N. All the results provided new insights into the key residues required for ACS and CSL activities of AtACS7 and laid an important foundation for further in-depth study of the biological functions of its dual enzymatic activities.
Funder
National Natural Science Foundation of China
Subject
Inorganic Chemistry,Organic Chemistry,Physical and Theoretical Chemistry,Computer Science Applications,Spectroscopy,Molecular Biology,General Medicine,Catalysis
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