Hindered rotation of a cofactor methyl group as a probe for protein-cofactor interaction
| Autor: | Tilo Mathes, Richard Brosi, Monika Joshi, Markus Fischer, Stefan Weber, Peter Hegemann, Robert Bittl, Adelbert Bacher, Boris Illarionov, Erik Schleicher |
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| Rok vydání: | 2010 |
| Předmět: |
Models
Molecular Phototropins biology Avena Rotation Stereochemistry Chemistry Flavin Mononucleotide Coenzymes Electron Spin Resonance Spectroscopy General Chemistry Biochemistry Catalysis Cofactor Protein Structure Tertiary chemistry.chemical_compound Colloid and Surface Chemistry Molecular level biology.protein Mutagenesis Site-Directed Chlamydomonas reinhardtii Methyl group Protein Binding |
| Zdroj: | Journal of the American Chemical Society. 132(26) |
| ISSN: | 1520-5126 |
| Popis: | Exploring protein-cofactor interactions on a molecular level is one of the major challenges in modern biophysics. Based on structural data alone it is rarely possible to identify how subtle interactions between a protein and its cofactor modulate the protein's reactivity. In the case of enzymatic processes in which paramagnetic molecules play a certain role, EPR and related methods such as ENDOR are suitable techniques to unravel such important details. In this contribution, we describe how cryogenic-temperature ENDOR spectroscopy can be applied to various LOV domains, the blue-light sensing domains of phototropin photoreceptors, to gain information on the direct vicinity of the flavin mononucleotide (FMN) cofactor by analyzing the temperature dependence of methyl-group rotation attached to C(8) of the FMN's isoalloxazine ring. More specifically, mutational studies of three amino acids surrounding the methyl group led to the identification of Asn425 as an important amino acid that critically influences the dark-state recovery of Avena sativa LOV2 domains. Consequently, it is possible to probe protein-cofactor interactions on a sub-angstrom level by following the temperature dependencies of hyperfine couplings. |
| Databáze: | OpenAIRE |
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