Abstract
ATP binding cassette (ABC) proteins are present in all phyla of life and form one of the largest protein families. The Bacillus subtilis ABC transporter BmrA is a functional homodimer that can extrude many different harmful compounds out of the cell. Each BmrA monomer is composed of a transmembrane domain (TMD) and a nucleotide binding domain (NBD). While the TMDs of ABC transporters are sequentially diverse, the highly conserved NBDs harbor distinctive conserved motifs that enable nucleotide binding and hydrolysis, interdomain communication and that mark a protein as a member of the ABC superfamily. In the catalytic cycle of an ABC transporter, the NBDs function as the molecular motor that fuels substrate translocation across the membrane via the TMDs and are thus pivotal for the entire transport process. For a better understanding of the structural and dynamic consequences of nucleotide interactions within the NBD at atomic resolution, we determined the 1H, 13C and 15N backbone chemical shift assignments of the 259 amino acid wildtype BmrA-NBD in its post-hydrolytic, ADP-bound state.
Highlights
In bacteria, the enhanced expression and production of efflux pumps can lead to multidrug resistance (MDR) which is aemerging problem for global human health
ATP binding cassette (ABC) transporters are an important family of MDR conferring membrane proteins and the only primary active transporters implicated in this phenomenon (Henderson et al 2021)
The molecular details of how ABC transporter nucleotide binding domain (NBD) sense nucleotides and allosterically mediate conformational changes remain under debate (Szöllősi et al 2018)
Summary
The enhanced expression and production of efflux pumps can lead to multidrug resistance (MDR) which is a (re)emerging problem for global human health. The molecular details of the local dynamic and structural consequences of nucleotide binding to the NBD as the central step of the ABC transporter catalytic cycle typically remain unresolved, either due to the use of site-specific labels or the significant signal overlap that such a large system evokes which typically precludes obtaining per residue information within the NBD upon nucleotide binding or hydrolysis Such issues can be circumvented with complementary solution NMR studies on the isolated NBDs. Importantly, the NBDs of ABC transporters are stable in isolation and maintain their structural and functional integrity, i.e. their ability to interact with nucleotides. We report the backbone resonance assignments of the isolated wildtype (WT) BmrA-NBD (residues G331-G589) in its post-hydrolysis, ADP-bound state as a model system for the detailed structural and dynamic analysis of the ABC transporter NBD as one of the most prominent functional domains found in all phyla of life
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