Physicochemical Characterisation of KEIF-The Intrinsically Disordered N-Terminal Region of Magnesium Transporter A.

Stéphanie Jephthah,Domagoj Belić,Marie Skepö,Jens Preben Morth,Linda K Månsson

doi:10.3390/biom10040623

Abstract

Magnesium transporter A (MgtA) is an active transporter responsible for importing magnesium ions into the cytoplasm of prokaryotic cells. This study focuses on the peptide corresponding to the intrinsically disordered N-terminal region of MgtA, referred to as KEIF. Primary-structure and bioinformatic analyses were performed, followed by studies of the undisturbed single chain using a combination of techniques including small-angle X-ray scattering, circular dichroism spectroscopy, and atomistic molecular-dynamics simulations. Moreover, interactions with large unilamellar vesicles were investigated by using dynamic light scattering, laser Doppler velocimetry, cryogenic transmission electron microscopy, and circular dichroism spectroscopy. KEIF was confirmed to be intrinsically disordered in aqueous solution, although extended and containing little β-structure and possibly PPII structure. An increase of helical content was observed in organic solvent, and a similar effect was also seen in aqueous solution containing anionic vesicles. Interactions of cationic KEIF with anionic vesicles led to the hypothesis that KEIF adsorbs to the vesicle surface through electrostatic and entropic driving forces. Considering this, there is a possibility that the biological role of KEIF is to anchor MgtA in the cell membrane, although further investigation is needed to confirm this hypothesis.

Highlights

In a society where antimicrobial resistance is constantly manifesting in new ways, the demand for effective antibiotics is naturally increasing
The primary structure was analysed to predict the overall structure and behaviour of the peptide. This was done by evaluating the charge, disorder propensity, and hydrophobicity/hydrophilicity per amino acid residue in the KEIF sequence
This paper featured the extensive physicochemical characterisation of KEIF, the N-terminal disordered region of Magnesium transporter A (MgtA), using an approach combining various experimental techniques and MD simulations. Both the experimental techniques and the complementary simulations confirmed that KEIF is an extended intrinsically disordered peptide with little propensity towards β-structures, and possibly polyproline II (PPII) structure

Summary

Introduction

In a society where antimicrobial resistance is constantly manifesting in new ways, the demand for effective antibiotics is naturally increasing. In order to rationalise the design of new antibiotics, and to find new potential cellular targets, bacterial biochemical functions must be mapped and fully understood. The magnesium ion Mg2+ is the most abundant divalent cation in any biological system and, it being an essential mineral nutrient and an absolute requirement for life, is present in every cell type in every living organism [1]. Mg2+ is an essential cofactor for more than enzymes, including important DNA and RNA polymerases; it is required for stabilisation of the ribosome–protein complex during protein synthesis. In adenosine triphosphate (ATP)-dependent enzymes, Mg2+ binds an ATP, the main unit of cellular energy, in the catalytic pocket, activating the phosphate ester towards hydrolysis [2]. In non-ATP-dependent enzymes, the role of Mg2+ is instead to hold a water molecule in a specific position, and this water molecule in turn helps to hold a particular structure in place or participates directly in the enzymatic reaction mechanism [2]

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