The MCR-3 inside linker appears as a facilitator of colistin resistance

Area of Science:

• Microbiology
• Molecular Biology
• Biochemistry

Background:

• Mobile colistin resistance (MCR) enzymes pose a significant public health threat.
• The molecular mechanisms underlying MCR enzyme function, particularly in conferring colistin resistance, are not fully understood.
• MCR enzymes belong to the lipid A-phosphoethanolamine (PEA) transferase family.

Purpose of the Study:

• To elucidate the molecular mechanism by which a functional variant of MCR-3 confers colistin resistance.
• To identify key structural or functional elements responsible for MCR-3 activity.
• To understand how genetic modifications lead to phenotypic colistin resistance.

Main Methods:

• Genetic analysis of Aeromonas species to identify functional MCR variants.
• Biochemical characterization of the MCR-3 variant (Ah762) and its components.
• Molecular dynamics (MD) simulations to study enzyme-substrate interactions.
• Isothermal titration calorimetry (ITC) to assess binding affinities and interactions.

Main Results:

• A functional MCR-3 variant (Ah762) was identified in certain Aeromonas species, arising from genetic duplication and amplification.
• A specific hinge linker, termed Linker 59, was identified as crucial for MCR-3 function.
• Linker 59 acts as a facilitator, enabling inactive MCR variants to regain colistin resistance.
• MD simulations and ITC revealed that Linker 59 facilitates the formation of a phosphatidylethanolamine (PE)-accessible pocket within MCR-3-like enzymes.

Conclusions:

• Linker 59 is an intrinsic facilitator within MCR-3 enzymes that is essential for conferring colistin resistance.
• The findings define a novel mechanism for MCR enzyme activation and function.
• This research provides critical insights into the molecular basis of mobile colistin resistance, aiding in the development of countermeasures.