ArticleJournal of the American Chemical Society2013
Detection of dihydrofolate reductase conformational change by FRET using two fluorescent amino acids.
Article in Journal of the American Chemical Society, 2013. The graph could read no effect estimate from its abstract, so it casts no vote on the map. Cited by 24 papers.
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Who cites it
24 citing papers in PubMed, 65 citations in OpenAlex.
- Expanding the fluorescence toolkit: molecular design, synthesis and biological application of unnatural amino acids.Chemical science · 2025Review
- One-Pot Synthesis of Alkynyl-Conjugated Phenylalanine Analogues for Peptide-Based Fluorescent Imaging.Organic letters · 2025Article
- Biological Regulation StudiedAccounts of chemical research · 2025Article
- Synthesis of β-Pyridyl α-Amino Acids: Conformationally Sensitive Charge Transfer-Based Fluorophores.Organic letters · 2024Article
- Synthesis and Photophysical Properties of Charge-Transfer-Based Pyrimidine-Derived α-Amino Acids.The Journal of organic chemistry · 2023Article
- Engineering of enzymes using non-natural amino acids.Bioscience reports · 2022Review
- Monitoring protein conformational changes using fluorescent nanoantennas.Nature methods · 2022Article
- Optimized incorporation of an unnatural fluorescent amino acid affords measurement of conformational dynamics governing high-fidelity DNA replication.The Journal of biological chemistry · 2020Article
- Facile synthesis of a novel genetically encodable fluorescent α-amino acid emitting greenish blue light.Chemical communications (Cambridge, England) · 2020Article
- Site-Specific Tryptophan Labels Reveal Local Microsecond-Millisecond Motions of Dihydrofolate Reductase.Molecules (Basel, Switzerland) · 2020Article
- An intrinsic FRET sensor of protein-ligand interactions.Organic & biomolecular chemistry · 2020Article
- Fluorescent amino acids as versatile building blocks for chemical biology.Nature reviews. Chemistry · 2020Review
- Synthesis of alanyl nucleobase amino acids and their incorporation into proteins.Bioorganic & medicinal chemistry · 2016Article
- Synthesis and Evaluation of a Library of Fluorescent Dipeptidomimetic Analogues as Substrates for Modified Bacterial Ribosomes.Biochemistry · 2016Article
- Ligand-Dependent Conformational Dynamics of Dihydrofolate Reductase.Biochemistry · 2016Article
- Comparison of strategies for non-perturbing labeling of α-synuclein to study amyloidogenesis.Organic & biomolecular chemistry · 2016Article
- Multiply labeling proteins for studies of folding and stability.Current opinion in chemical biology · 2015Review
- Ribosome-Mediated Incorporation of Dipeptides and Dipeptide Analogues into Proteins in Vitro.Journal of the American Chemical Society · 2015Article
- Homologous ligands accommodated by discrete conformations of a buried cavity.Proceedings of the National Academy of Sciences of the United States of America · 2015Article
- Tyrosine-derived stimuli responsive, fluorescent amino acids.Chemical science · 2015Article
Corrections and comments
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Authors and funding
6 authors at 3 institutions in 1 country.
Funding
Abstract
Two fluorescent amino acids, including the novel fluorescent species 4-biphenyl-l-phenylalanine (1), have been incorporated at positions 17 and 115 of dihydrofolate reductase (DHFR) to enable a study of conformational changes associated with inhibitor binding. Unlike most studies involving fluorescently labeled proteins, the fluorophores were incorporated into the amino acid side chains, and both probes [1 and L-(7-hydroxycoumarin-4-yl)ethylglycine (2)] were smaller than fluorophores typically used for such studies. The DHFR positions were chosen as potentially useful for Förster resonance energy transfer (FRET) measurements on the basis of their estimated separation (17-18 Å) and the expected change in distance along the reaction coordinate. Also of interest was the steric accessibility of the two sites: Glu17 is on the surface of DHFR, while Ile115 is within a folded region of the protein. Modified DHFR I (1 at position 17; 2 at position 115) and DHFR II (2 at position 17; 1 at position 115) were both catalytically competent. However, DHFR II containing the potentially rotatable biphenylphenylalanine moiety at sterically encumbered position 115 was significantly more active than DHFR I. Irradiation of the modified DHFRs at 280 nm effected excitation of 1, energy transfer to 2, and emission by 2 at 450 nm. However, the energy transfer was substantially more efficient in DHFR II. The effect of inhibitor binding was also measured. Trimethoprim mediated concentration-dependent diminution of the emission observed at 450 nm for DHFR II but not for DHFR I. These findings demonstrate that amino acids containing small fluorophores can be introduced into DHFR with minimal disruption of function and in a fashion that enables sensitive monitoring of changes in DHFR conformation.
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