Crystal structure of a catalytic intermediate of the maltose transporter

Abstract

The maltose uptake system of Escherichia coli is a well-characterized member of the ATP-binding cassette transporter superfamily. Here we present the 2.8-A crystal structure of the intact maltose transporter in complex with the maltose-binding protein, maltose and ATP. This structure, stabilized by a mutation that prevents ATP hydrolysis, captures the ATP-binding cassette dimer in a closed, ATP-bound conformation. Maltose is occluded within a solvent-filled cavity at the interface of the two transmembrane subunits, about halfway into the lipid bilayer. The binding protein docks onto the entrance of the cavity in an open conformation and serves as a cap to ensure unidirectional translocation of the sugar molecule. These results provide direct evidence for a concerted mechanism of transport in which solute is transferred from the binding protein to the transmembrane subunits when the cassette dimer closes to hydrolyse ATP.

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).…”

“…Although the hinge motion described by this transformation is a mathematical construct that need not represent the actual motion, it is intriguing that the rotation axis passes close to the conserved Pro67 of both subunits found at the kink on the periplasmic side of TM2; this region corresponds to the periplasmic gate identified in the MalFGK 2 and ModBC structures, and further is adjacent to the internal binding site for maltose found in the MalFGK 2 structure. 13 The rotation axis corresponding to the rigid body change between the TMD and NBD also passes through elements conserved in Type I importers, including the coupling helices connecting the TMDs and NBDs.…”

“…The structurally equivalent residues to Met163 in the maltose transporter are Val442 and Val230 of MalF and MalG, respectively. 13 In the methionine transporter, the Met163 side chains from opposing subunits pack together in a gate like configuration [ Fig. 2(B)].…”

Structural and mechanistic insights into prokaryotic energy-coupling factor transporters

“…In contrast to the ATPases, the structures of the integral membrane subunits are not universally conserved among ABC transporters. Four different structural classes have been discovered, which probably have different mechanisms of substrate translocation (see the figure): type I importers (which are exemplified by the maltose transporter MalEFGK 2 from Escherichia coli [57][58][59][60][61] ; Protein Data Bank (PDB) accession 2R6G); type II importers (exemplified by vitamin B 12 transporter BtuC 2 D 2 F from E. coli [62][63][64] ; PDB accession 4FI3); exporters (such as multidrug and peptide transporters [65][66][67] , exemplified by the drug exporter TM287/288) from Thermotoga maritima; PDB accession 3QF4); and ECF transporters (such as ECF-FolT from Lactobacillus brevis for folate transport 21 ; PDB accession 4HUQ). Type I and II importers and energy-coupling factor (ECF) transporters are only found in prokaryotes, whereas exporters are found in all kingdoms of life 28 .…”

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.…”