Expulsion mechanism of the substrate-translocating subunit in ECF transporters

Abstract

Energy-coupling factor (ECF)-type transporters mediate the uptake of micronutrients in many bacteria. They consist of a substrate-translocating subunit (S-component) and an ATP-hydrolysing motor (ECF module) Previous data indicate that the S-component topples within the membrane to alternately expose the binding site to either side of the membrane. In many ECF transporters, the substrate-free S-component can be expelled from the ECF module. Here we study this enigmatic expulsion step by cryogenic electron microscopy and reveal that ATP induces a concave-to-convex shape change of two long helices in the motor, thereby destroying the S-component’s docking site and allowing for its dissociation. We show that adaptation of the membrane morphology to the conformational state of the motor may favour expulsion of the substrate-free S-component when ATP is bound and docking of the substrate-loaded S-component after hydrolysis. Our work provides a picture of bilayer-assisted chemo-mechanical coupling in the transport cycle of ECF transporters.

Coenzyme A biosynthesis inBacillus subtilis: Discovery of a novel precursor metabolite for salvage and its uptake system

“…The EcfA1 and EcfA2 subunits additionally interact with each other via their C-terminal domains (see http://www.subtiwiki.unigoettingen.de/v4/predictedComplex?id=173 for an interactive presentation of the complex). The model is in excellent agreement with the cryo-electron microscopic structure of the folate ECT transporter of Lactobacillus delbrückii (22).…”

The S-component fold: a link between bacterial transporters and receptors

“…S-components from ECF transporters are specialized in binding their substrate with high affinity, and enable the translocation of substrate upon the association with a tripartite ATPase ECF module 27 , 28 . The association leads to a characteristic toppling motion in which the S-component rotates relatively to the membrane plane, which brings the substrate binding site from an outward- to an inward-facing orientation, allowing substrate release into the cytoplasm.…”

“…ECF transporters consist of a three-subunit motor module (termed ECF module) that binds and hydrolyzes ATP, and an integral membrane subunit (S-component or EcfS) that binds and translocates the cognate substrate across the membrane upon association with the motor module (Fig. 1 ) 27 , 28 . While the ECF module is well conserved with multiple typifying motifs (i.e., Walker A and B motifs, signature motifs of the ATPase subunits, and Ala-Arg-Gly motifs of the scaffold protein EcfT) 26 , 29 , S-components with specificity for different transported substrates display highly variable sequences, which has complicated the identification of the encoding genes.…”

The S-component fold: A link between bacterial transporters and receptors

“…S-components from ECF transporters are specialized in binding their substrate with high affinity, and enable the translocation of substrate upon the association with a tripartite ATPase ECF-module. 64,65 The association leads to a characteristic toppling motion in which the S-component rotates relatively to the membrane plane, which brings the substrate binding site from an outward- to an inward-facing orientation, allowing substrate release into the cytoplasm. The assembly of the S-component and the ECF module into a full ECF transporter (Figure 5A) involves an AXXXA motif in helix 1, which is relatively conserved among S-components, as evidenced by the structures of the full complexes determined to date.…”