ArticleProceedings of the National Academy of Sciences of the United States of America2000
Redesign of substrate-selectivity determining modules of glutathione transferase A1-1 installs high catalytic efficiency with toxic alkenal products of lipid peroxidation.
Article in Proceedings of the National Academy of Sciences of the United States of America, 2000. The graph could read no effect estimate from its abstract, so it casts no vote on the map. Cited by 26 papers.
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26 citing papers in PubMed, 79 citations in OpenAlex.
- Human mitochondrial glutathione transferases: Kinetic parameters and accommodation of a mitochondria-targeting group in substrates.Bioorganic & medicinal chemistry · 2024Article
- Reversibility and Low Commitment to Forward Catalysis in the Conjugation of Lipid Alkenals by Glutathione Transferase A4-4.Biomolecules · 2023Article
- Glutathione Transferases as Efficient Ketosteroid Isomerases.Frontiers in molecular biosciences · 2021Review
- Water-Soluble Pristine CPharmaceutics · 2020Article
- Substrate structure and computation guided engineering of a lipase for omega-3 fatty acid selectivity.PloS one · 2020Article
- Water-Soluble Pristine COxidative medicine and cellular longevity · 2020Article
- Exploring sequence-function space of a poplar glutathione transferase using designed information-rich gene variants.Protein engineering, design & selection : PEDS · 2017Article
- Drosophila GSTs display outstanding catalytic efficiencies with the environmental pollutants 2,4,6-trinitrotoluene and 2,4-dinitrotoluene.Biochemistry and biophysics reports · 2016Article
- Role of CRD-BP in the growth of human basal cell carcinoma cells.The Journal of investigative dermatology · 2014Article
- Enzymatic detoxication, conformational selection, and the role of molten globule active sites.The Journal of biological chemistry · 2013Article
- Cloning, expression and analysis of the olfactory glutathione S-transferases in coho salmon.Biochemical pharmacology · 2013Article
- Ensemble perspective for catalytic promiscuity: calorimetric analysis of the active site conformational landscape of a detoxification enzyme.The Journal of biological chemistry · 2011Article
- Cys-X scanning for expansion of active-site residues and modulation of catalytic functions in a glutathione transferase.The Journal of biological chemistry · 2011Article
- Interactions of glutathione transferases with 4-hydroxynonenal.Drug metabolism reviews · 2011Review
- Substrate specificity combined with stereopromiscuity in glutathione transferase A4-4-dependent metabolism of 4-hydroxynonenal.Biochemistry · 2010Article
- Minor modifications of the C-terminal helix reschedule the favored chemical reactions catalyzed by theta class glutathione transferase T1-1.The Journal of biological chemistry · 2010Article
- The human hGSTA5 gene encodes an enzymatically active protein.Biochimica et biophysica acta · 2010Article
- Structural analysis of a glutathione transferase A1-1 mutant tailored for high catalytic efficiency with toxic alkenals.Biochemistry · 2009Article
- The stereochemical course of 4-hydroxy-2-nonenal metabolism by glutathione S-transferases.The Journal of biological chemistry · 2008Article
- The intersubunit lock-and-key motif in human glutathione transferase A1-1: role of the key residues Met51 and Phe52 in function and dimer stability.The Biochemical journal · 2006Article
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3 authors at 1 institution in 1 country.
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No grant is acknowledged in the PubMed record.
Abstract
The evolution of proteins for novel functions involves point mutations and recombinations of domains or structural segments. Mimicking this process by rational design in vitro is still a major challenge. The present report demonstrates that the active site of the enzyme glutathione transferase (GST) A1-1 can be tailored for high catalytic efficiency with alkenals. The result is a >3,000-fold change in substrate selectivity involving a noteworthy change in preferred catalyzed reaction from aromatic nucleophilic substitution to Michael addition. The hydrophobic substrate binding pocket of GST A1-1 is formed by three structural modules, which were redesigned sequentially with four point mutations and the exchange of a helical segment. The substitutions were made to mimic first-sphere interactions with a substrate in GST A4-4, which naturally has high activity with alkenals. These substrates are toxic lipid peroxidation products of pathophysiological significance, and glutathione conjugation is a route of their inactivation. The final product of the sequential redesign of GST A1-1, mutant GIMFhelix, had a 300-fold increase in catalytic efficiency with nonenal and a >10 times decreased activity with 1-chloro-2,4-dinitrobenzene. In absolute values, GIMFhelix is more efficient than wild-type GST A4-4 with some alkenal substrates, with a k(cat)/K(m) value of 1.5 +/- 0. 1 10(6) M(-1) small middle dots(-1) for nonenal. The pKa value of the active-site Tyr-9 of GIMFhelix is 7.3 +/- 0.1, approaching the unusually low value of GST A4-4. Thus, rational redesign of the active-site region of an enzyme may be sufficient for the generation of efficient catalysts with altered chemical mechanism and novel selectivity.
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