9u2x | pdb_00009u2x
Crystal structure of N244L variant from Bacillus subtilis
Structural highlights
FunctionEFEB_BACSU Involved in the recovery of exogenous heme iron (By similarity). Extracts iron from heme while preserving the protoporphyrin ring intact (By similarity). Part of the iron transporter system efeUOB/M involved in iron import (PubMed:16672620, PubMed:23764491). Catalyzes the peroxide-mediated oxidation of Fe(2+) into Fe(3+); EfeM binds Fe(3+) and delivers it to the cell membrane permease EfeU (PubMed:23764491).[UniProtKB:P31545][1] [2] Publication Abstract from PubMedDye-decolorizing peroxidases (DyPs) are heme enzymes with broad substrate scope and increasing biotechnological relevance, yet the structural determinants governing their activation by reactive oxygen species (ROS) remain incompletely understood. Here, we identify dynamic ROS gating as a mechanism controlling activation and catalytic efficiency through a comparative study of two class I DyPs from Bacillus subtilis (BsDyP) and Thermobifida fusca (TfuDyP), together with the BsDyP N244L variant. By combining steady-state kinetics, ROS-selective electroreductive activation assays, UV-vis and resonance Raman spectroscopy, X-ray crystallography, and molecular dynamics simulations, we establish a direct structure-function relationship linking distal heme pocket organization and access-tunnel architecture to enzyme-specific ROS preferences. BsDyP WT is preferentially activated via (*)OH, whereas TfuDyP relies mainly on H(2)O(2). Remarkably, the single N244L substitution shifts the catalytic, structural, and dynamical properties of BsDyP toward those of TfuDyP, making both enzymes nearly indistinguishable in ROS usage and catalytic efficiency. This convergence arises from a reorganization of distal hydrogen-bonding networks and loop-mediated reshaping of the access tunnels, which together bias ROS accessibility and lower the barrier for compound I formation. These findings establish ROS gating as the principal determinant of DyP activation and provide a mechanistic framework for tuning ROS selectivity through protein engineering. A Single Amino Acid Substitution Reprograms ROS Selectivity and Catalytic Function in DyP Peroxidases.,Zitare UA, Castro MA, Scocozza MF, Borges PT, Martins LO, Murgida DH Inorg Chem. 2026 Jun 15. doi: 10.1021/acs.inorgchem.6c02237. PMID:42295056[3] From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine. References
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