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Catalytic mechanism of the colistin resistance protein MCR-1
Reynier Suardíaz1, Emily Lythell2, Philip Hinchliffe3
1Centre for Computational Chemistry, School of Chemistry, University of Bristol, Cantock's Close, Bristol BS8 1TS, UK. reysuard@ucm.es adrian.mulholland@bristol.ac.uk and School of Biochemistry, University of Bristol, University Walk, Bristol BS8 1TD, UK and Departamento de Química Física, Facultad de Química, Universidad Complutense, 28040 Madrid, Spain.
Abstract:
The mcr-1 gene encodes a membrane-bound Zn2+-metalloenzyme, MCR-1, which catalyses phosphoethanolamine transfer onto bacterial lipid A, making bacteria resistant to colistin, a last-resort antibiotic. Mechanistic understanding of this process remains incomplete. Here, we investigate possible catalytic pathways using DFT and ab initio calculations on cluster models and identify a complete two-step reaction mechanism. The first step, formation of a covalent phosphointermediate via transfer of phosphoethanolamine from a membrane phospholipid donor to the acceptor Thr285, is rate-limiting and proceeds with a single Zn2+ ion. The second step, transfer of the phosphoethanolamine group to lipid A, requires an additional Zn2+. The calculations suggest the involvement of the Zn2+ orbitals directly in the reaction is limited, with the second Zn2+ acting to bind incoming lipid A and direct phosphoethanolamine addition. The new level of mechanistic detail obtained here, which distinguishes these enzymes from other phosphotransferases, will aid in the development of inhibitors specific to MCR-1 and related bacterial phosphoethanolamine transferases.
Insights
The MCR-1 enzyme confers colistin resistance by transferring phosphoethanolamine to bacterial lipid A. Computational studies reveal a two-step mechanism, crucial for developing new MCR-1 inhibitors.
Area of Science:
- Biochemistry
- Computational Chemistry
- Antimicrobial Resistance
Background:
- The mcr-1 gene encodes the MCR-1 enzyme, a Zn2+-metalloenzyme.
- MCR-1 mediates phosphoethanolamine transfer to bacterial lipid A, conferring resistance to colistin.
- Understanding the MCR-1 catalytic mechanism is incomplete.
Purpose of the Study:
- To elucidate the complete catalytic mechanism of MCR-1 using computational methods.
- To identify key steps and metal ion involvement in phosphoethanolamine transfer.
- To provide mechanistic insights for developing MCR-1 specific inhibitors.
Main Methods:
- Density Functional Theory (DFT) calculations.
- Ab initio calculations on cluster models.
- Investigation of reaction pathways for phosphoethanolamine transfer.
Main Results:
- A complete two-step reaction mechanism for MCR-1 was identified.
- The first step involves a rate-limiting covalent phosphointermediate formation with one Zn2+ ion.
- The second step requires a second Zn2+ ion to bind lipid A and direct transfer.
Conclusions:
- The study provides unprecedented mechanistic detail of MCR-1 activity.
- The findings distinguish MCR-1 from other phosphotransferases.
- This knowledge will facilitate the design of targeted inhibitors against MCR-1 and related enzymes.
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