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Active Site Aromatic Residues Play a Dual Role in the Substrate Interaction and Protein Structure in Functional Dimers of CYP121A1 of Mycobacterium tuberculosis

The essential enzyme CYP121A1 of Mycobacterium tuberculosis forms a functional dimer, which when disrupted results in a decrease of activity and substrate specificity. The crystal structure of CYP121A1 in complex with its substrate di-cyclotyrosine (cYY) indicates that the aromatic side chains of Ph...

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Bibliographic Details
Published in:ACS infectious diseases 2023-04, Vol.9 (4), p.827-839
Main Authors: Campomizzi, Christopher S., Kumar, Amit, Uttamrao, Patil Pranita, Stallone, Jack J., Ghanatios, George E., Rathinavelan, Thenmalarchelvi, Estrada, D. Fernando
Format: Article
Language:English
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Summary:The essential enzyme CYP121A1 of Mycobacterium tuberculosis forms a functional dimer, which when disrupted results in a decrease of activity and substrate specificity. The crystal structure of CYP121A1 in complex with its substrate di-cyclotyrosine (cYY) indicates that the aromatic side chains of Phe-168 and Trp-182 form stabilizing π–π interactions with a tyrosyl ring of cYY. In the enclosed study, we utilize targeted 19F labeling of aromatic residues to label CYP121A1 for detection by nuclear magnetic resonance (NMR) spectroscopy. 19F-NMR spectra and functional characterization of mutations to Phe-168 and Trp-182 are combined with all-atom molecular dynamics simulations of substrate-bound and substrate-free CYP121A1. This study shows that these aromatic residues interact with cYY predominantly through π–π stacking. In addition to playing an essential role in substrate binding, these active site residues also stabilize the tertiary and quaternary structures of CYP121A1. An additional unexpected finding was the presence of cYY-induced long-range allostery that affects residues located near the homodimer interface. Taken together, this study highlights a structural relationship between the active site environment of this essential enzyme with its global structure that was previously unknown.
ISSN:2373-8227
2373-8227
DOI:10.1021/acsinfecdis.2c00531