Abstract
ATP-binding cassette (ABC) type I importers are widespread in bacteria and play a crucial role in its survival and pathogenesis. They share the same modular architecture comprising two intracellular nucleotide-binding domains (NBDs), two transmembrane domains (TMDs) and a substrate-binding protein. The NBDs bind and hydrolyze ATP, thereby generating conformational changes that are coupled to the TMDs and lead to substrate translocation. A group of multitask NBDs that are able to serve as the cellular motor for multiple sugar importers was recently discovered. To understand why some ABC importers share energy-coupling components, we used the MsmX ATPase from Bacillus subtilis as a model for biological and structural studies. Here we report the first examples of functional hybrid interspecies ABC type I importers in which the NBDs could be exchanged. Furthermore, the first crystal structure of an assigned multitask NBD provides a framework to understand the molecular basis of the broader specificity of interaction with the TMDs.
Highlights
ATP-binding cassette (ABC) transporters are found in all three domains of life, from bacteria to humans, and constitute a large superfamily of membrane translocator proteins of a wide variety of molecules, such as ions, amino acids, sugars, lipids, peptides, proteins and antibiotics[1,2]
To date, proteins with this ability have been exclusively reported in bacterial ABC sugar importers of the subfamily of carbohydrate uptake transporter 1 (CUT1), that transport a variety of di, tri‐ and higher oligosaccharides, as well as polyols[21,22]
To characterize the energy-coupling component of bacterial ABC transporters, and to evaluate their intra- and interspecies exchangeability, we constructed a genetic system in B. subtilis for regulated expression of the msmX allele or its homologs in trans (Fig. 1)
Summary
ATP-binding cassette (ABC) transporters are found in all three domains of life, from bacteria to humans, and constitute a large superfamily of membrane translocator proteins of a wide variety of molecules, such as ions, amino acids, sugars, lipids, peptides, proteins and antibiotics[1,2]. The two groups of transporters share the same NBD fold but their TMDs are totally different: the two TMDs of type I importers are either identical or structurally similar (like the prototype maltose transporter M alEFGK2 from Escherichia coli10), with a core membrane topology of five TM helices per TMD, while those of type II have two identical TMDs each containing ten TM helices. Other multitask ABC ATPases have been identified in other bacteria such as Streptomyces lividans[19] and Streptococcus suis[20], the ability to interact with and energize multiple transporters is rather uncommon among ABC NBDs. to date, proteins with this ability have been exclusively reported in bacterial ABC sugar importers of the subfamily of carbohydrate uptake transporter 1 (CUT1), that transport a variety of di‐, tri‐ and higher oligosaccharides, as well as polyols[21,22]. We solve the crystal structure of the B. subtilis multitask ATPase MsmX K43A; the crystals diffracted up to 1.67 Å resolution and based on comparative analysis we hypothesize about sequence and structural features involved in the broader specificity of NBDs
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