The structural basis of modularity in ECF-type ABC transporters

Abstract

Energy coupling factor (ECF) transporters are used for the uptake of vitamins in Prokarya. They consist of an integral membrane protein that confers substrate specificity (the S-component) and an energizing module that is related to ATP-binding cassette (ABC) transporters. S-components for different substrates often do not share detectable sequence similarity but interact with the same energizing module. Here we present the crystal structure of the thiamine-specific S-component ThiT from Lactococcus lactis at 2.0 Å. Extensive protein-substrate interactions explain its high binding affinity for thiamine (K(d) ~10(-10) M). ThiT has a fold similar to that of the riboflavin-specific S-component RibU, with which it shares only 14% sequence identity. Two alanines in a conserved motif (AxxxA) located on the membrane-embedded surface of the S-components mediate the interaction with the energizing module. Based on these findings, we propose a general transport mechanism for ECF transporters.

Structural and mechanistic insights into prokaryotic energy-coupling factor transporters

“…Molecular dynamics simulations of the solitary S component for thiamin ThiT, of which a crystal structure had been determined before the structures of the complete ECF transporters, are consistent with these generic features of membrane proteins 25,39 (FIG. 2b).…”

“…Both of these metal-binding S components seem to require auxiliary small integral membrane proteins for substrate binding -CbiN for CbiM, and either NikN or NikKL (which is a complex of two proteins) for NikM 6,69,70 -but the exact role of these additional proteins remains to be elucidated. CbiM and NikM have seven predicted membrane-spanning α-helices, in contrast to the common six helices of other S components [23][24][25] . Whether CbiM and NikM share the conserved structural core of the other S components is not clear.…”

“…Unexpectedly, the orientation of the S components FolT and HmpT in the complete ECF transporters is entirely different, even though their global folds are the same as RibU, ThiT and BioY [21][22][23][24][25] (FIG. 2c).…”

“…So far, 21 different S component families have been identified, each of which is specific for a distinct substrate 3,4,6 (TABLE 1). Although S components for different substrates lack a high degree of sequence similarity (they have on average 10-20% sequence identity), crystal structures of isolated S components have shown that their global structures are conserved [23][24][25] . It is likely that this fold is shared by all S components (with the possible exception of the S components for Co 2+ and Ni 2+ ; BOX 1).…”

Dynamic Combinatorial Chemistry to Identify Binders of ThiT, an S‐Component of the Energy‐Coupling Factor Transporter for Thiamine

“…Up to date, there are only a few reports in which DCC has been applied to identify binders of transmembrane proteins 4, 5, 6. Here, we have applied DCC to identify possible binders for ThiT, the S‐component of the energy‐coupling factor (ECF) transporter for thiamine (Figure 1 A) in Lactococcus lactis 7, 8. ECF transporters represent an interesting target for the development of antibacterial agents with a novel mode of action by blocking vitamin transport 9…”