The Soybean Lipoxygenase-Substrate Complex: Correlation Between Properties of Tunneling-Ready States and ENDOR-Detected Structures of Ground States
| Autor: | Judith P. Klinman, Ajay Sharma, Adam R. Offenbacher, Peter E. Doan, Brian M. Hoffman |
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| Jazyk: | angličtina |
| Rok vydání: | 2020 |
| Předmět: |
Hydrogen
Protein Conformation Population Lipoxygenase chemistry.chemical_element Biochemistry Article Linoleic Acid 03 medical and health sciences chemistry.chemical_compound symbols.namesake Side chain Carbon-13 Magnetic Resonance Spectroscopy education Conformational isomerism Quantum tunnelling 0303 health sciences education.field_of_study Carbon Isotopes biology Chemistry 030302 biochemistry & molecular biology Active site Crystallography Mutation symbols biology.protein Hydroxide Soybeans van der Waals force |
| Zdroj: | Biochemistry |
| Popis: | Hydrogen tunneling in enzymatic C-H activation requires a dynamical sampling among ground-state enzyme-substrate (E-S) conformations, which transiently generates a tunneling-ready state (TRS). The TRS is characterized by a hydrogen donor-acceptor distance (DAD) of 2.7 A, ∼0.5 A shorter than the dominant DAD of optimized ground states. Recently, a high-resolution, 13C electron-nuclear double-resonance (ENDOR) approach was developed to characterize the ground-state structure of the complex of the linoleic acid (LA) substrate with soybean lipoxygenase (SLO). The resulting enzyme-substrate model revealed two ground-state conformers with different distances between the target C11 of LA and the catalytically active cofactor [Fe(III)-OH]: the active conformer "a", with a van der Waals DAD of 3.1 A between C11 and metal-bound hydroxide, and an inactive conformer "b", with a distance that is almost 1 A longer. Herein, the structure of the E-S complex is examined for a series of six variants in which subtle structural modifications of SLO have been introduced either at a hydrophobic side chain near the bound substrate or at a remote residue within a protein network whose flexibility influences hydrogen transfer. A remarkable correlation is found between the ENDOR-derived population of the active ground-state conformer a and the kinetically derived differential enthalpic barrier for D versus H transfer, ΔEa, with the latter increasing as the fraction of conformer a decreases. As proposed, ΔEa provides a "ruler" for the DAD within the TRS. ENDOR measurements further corroborate the previous identification of a dynamical network coupling the buried active site of SLO to the surface. This study shows that subtle imperfections within the initial ground-state structures of E-S complexes are accompanied by compromised geometries at the TRS. |
| Databáze: | OpenAIRE |
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