A Key Role in Catalysis and Enzyme Thermostability of a Conserved Helix H5 Motif of Human Glutathione Transferase A1-1
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Published:2023-02-12
Issue:4
Volume:24
Page:3700
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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:
Chronopoulou Evangelia G.1ORCID, Mutabdzija Lana2, Poudel Nirmal2, Papageorgiou Anastassios C.2ORCID, Labrou Nikolaos E.1ORCID
Affiliation:
1. Laboratory of Enzyme Technology, Department of Biotechnology, School of Applied Biology and Biotechnology, Agricultural University of Athens, 75 Iera Odos Street, 11855 Athens, Greece 2. Turku Bioscience Centre, University of Turku and Åbo Akademi University, 20520 Turku, Finland
Abstract
Glutathione transferases (GSTs) are promiscuous enzymes whose main function is the detoxification of electrophilic compounds. These enzymes are characterized by structural modularity that underpins their exploitation as dynamic scaffolds for engineering enzyme variants, with customized catalytic and structural properties. In the present work, multiple sequence alignment of the alpha class GSTs allowed the identification of three conserved residues (E137, K141, and S142) at α-helix 5 (H5). A motif-directed redesign of the human glutathione transferase A1-1 (hGSTA1-1) was performed through site-directed mutagenesis at these sites, creating two single- and two double-point mutants (E137H, K141H, K141H/S142H, and E137H/K141H). The results showed that all the enzyme variants displayed enhanced catalytic activity compared to the wild-type enzyme hGSTA1-1, while the double mutant hGSTA1-K141H/S142H also showed improved thermal stability. X-ray crystallographic analysis revealed the molecular basis of the effects of double mutations on enzyme stability and catalysis. The biochemical and structural analysis presented here will contribute to a deeper understanding of the structure and function of alpha class GSTs.
Funder
ΙΚΥ Scholarship Programs Strengthening PostDoctoral Research Operational Programme “Human Resources Development Program, Education and Lifelong Learning” European Social Fund—ESF and the Greek government Biocenter Finland and Academy of Finland Magnus Ehrnrooth Foundation Horizon 2020 programme of the European Union
Subject
Inorganic Chemistry,Organic Chemistry,Physical and Theoretical Chemistry,Computer Science Applications,Spectroscopy,Molecular Biology,General Medicine,Catalysis
Reference51 articles.
1. Recent advances in protein engineering and biotechnological applications of glutathione transferases;Perperopoulou;Crit. Rev. Biotechnol.,2018 2. Glutathione transferases: Substrates, inihibitors and pro-drugs in cancer and neurodegenerative diseases;Allocati;Oncogenesis,2018 3. Erythrocyte glutathione transferase: A general probe for chemical contaminations in mammals;Bocedi;Cell Death Discov.,2016 4. Fabrini, R., Bocedi, A., Pallottini, V., Canuti, L., De Canio, M., Urbani, A., Marzano, V., Cornetta, T., Stano, P., and Giovanetti, A. (2010). Nuclear shield. A multi-enzyme task- force for nucleus protection. PLoS ONE, 5. 5. The impact of nitric oxide toxicity on the evolution of the glutathione transferase superfamily: A proposal for an evolutionary driving force;Bocedi;J. Biol. Chem.,2013
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