Structure of an ABC transporter in complex with its binding protein

Abstract

ATP-binding cassette (ABC) transporter proteins carry diverse substrates across cell membranes. Whereas clinically relevant ABC exporters are implicated in various diseases or cause multidrug resistance of cancer cells, bacterial ABC importers are essential for the uptake of nutrients, including rare elements such as molybdenum. A detailed understanding of their mechanisms requires direct visualization at high resolution and in distinct conformations. Our recent structure of the multidrug ABC exporter Sav1866 has revealed an outward-facing conformation of the transmembrane domains coupled to a closed conformation of the nucleotide-binding domains, reflecting the ATP-bound state. Here we present the 3.1 A crystal structure of a putative molybdate transporter (ModB2C2) from Archaeoglobus fulgidus in complex with its binding protein (ModA). Twelve transmembrane helices of the ModB subunits provide an inward-facing conformation, with a closed gate near the external membrane boundary. The ATP-hydrolysing ModC subunits reveal a nucleotide-free, open conformation, whereas the attached binding protein aligns the substrate-binding cleft with the entrance to the presumed translocation pathway. Structural comparison of ModB2C2A with Sav1866 suggests a common alternating access and release mechanism, with binding of ATP promoting an outward-facing conformation and dissociation of the hydrolysis products promoting an inward-facing conformation.

Inward facing conformations of the MetNI methionine ABC transporter: Implications for the mechanism of transinhibition

“…The structural foundations for this analysis are most advanced for Type I importers, 10 particularly for the maltose MalFGK 2 transporter, [25][26][27][28] and provide an important framework for interpreting the mechanistic significance of the current MetNI structures. Structurally characterized type I importers include MalFGK 2 in outward facing (PDB IDs 2R6G 13 and 3PUY 27 ), pretranslocation (PDB IDs 3PV0 and 3PUZ 27 ), and inward facing (3FH6 36 ) conformations, ModABC in the inward and inward/inhibited conformations (PDB IDs 2ONK 24 and 3D31, 32 respectively), and the CY5 and DM forms of MetNI (PDB IDs 3TUI and 3TUJ, respectively) presented in this work. Although the TMDs of Type I importers are organized around a common core of five TM helices, a subset of three of these helices, TM2-4 in MetNI, are highly conserved and exhibit a one to one residue correspondence that can be used for quantitative comparisons of the relationships of the TMDs between different structures; a conserved core can also be identified for the NBDs (that is, of course, general to the entire ABC transporter superfamily).…”

“…[19][20][21][22] In the conformation competent for ATP hydrolysis, the nucleotides are sandwiched between the conserved P-loop and ABC signature motifs from opposing ABC subunits. The conformational changes accompanying nucleotide binding and hydrolysis are then transmitted to the membrane spanning domains through coupling helices 23,24 present in cytoplasmic loops between transmembrane helices. The mechanistic understanding of the transporter cycle is most advanced for Type I importers, particularly the maltose MalFGK 2 importer, where a combination of biochemical, biophysical, and structural studies have provided important insights.…”

Dynamics and Structural Changes Induced by ATP Binding in SAV1866, a Bacterial ABC Exporter

“…All but one correspond to bacterial transporters. Five are ABC importers (BtuCD, ModBC-A, HI1470/1, MalFG/ K, MetNI), [9][10][11][12][13][14][15] and three are drug exporters (SAV1866, MsbA, and mouse P-glycoprotein). [16][17][18][19] High-resolution structures and NMR, EPR, and electron microscopy data strongly suggest that ABC exporters undergo large movements during substrate efflux, in particular upon ATP binding and/or hydrolysis.…”

cAMP-dependent Protein Kinase Phosphorylation Produces Interdomain Movement in SUR2B Leading to Activation of the Vascular KATP Channel

“…Several ABC proteins have been crystallized in full-length with TMDs (33)(34)(35)(36). These include the recently crystallized SAV1866 that is a homologue of mammalian ABC proteins (36) and shows the highest sequence similarity to SUR2B.…”