Uracil/H+ symport by the FurE transporter challenges the rocking-bundle mechanism of transport in APC transporters

Zantza, Iliana; Papadaki, Georgia F.; Raniolo, Stefano; Pyrris, Yiannis; Lambrinidis, George; Limongelli, Vittorio; Diallinas, George; Mikros, Emmanuel

doi:10.1101/2022.03.28.486045

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Uracil/H+ symport by the FurE transporter challenges the rocking-bundle mechanism of transport in APC transporters

Zantza, Iliana Department of Pharmacy, National and Kapodistrian University of Athens, Greece
Papadaki, Georgia F. Department of Biology, National and Kapodistrian University of Athens, Greece
Raniolo, Stefano ORCID Euler Institute (EUL), Università della Svizzera italiana, Switzerland
Pyrris, Yiannis Department of Biology, National and Kapodistrian University of Athens, Greece
Lambrinidis, George Department of Pharmacy, National and Kapodistrian University of Athens, Greece
Limongelli, Vittorio ORCID Euler Institute (EUL), Università della Svizzera italiana, Switzerland - Department of Pharmacy, University of Naples “Federico II”, Italy
Diallinas, George Department of Biology, National and Kapodistrian University of Athens, Greece - Institute of Molecular Biology and Biotechnology, Foundation for Research and Technology, Heraklion, Greece
Mikros, Emmanuel Department of Pharmacy, National and Kapodistrian University of Athens, Greece - Athena Research and Innovation Center in Information Communication & Knowledge Technologies, Marousi, Greece

2022

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Journal of molecular biology. - 2022, vol. 435, no. 19, p. 168226

English Transporters mediate the uptake of solutes, metabolites and drugs across the cell membrane. The eukaryotic FurE nucleobase/H+ symporter of Aspergillus nidulans has been used as a model protein to address structure-function relationships in the APC transporter superfamily, members of which are characterized by the LeuT-fold and seem to operate by the so-called ‘rocking-bundle’ mechanism. In this study, we reveal the binding mode, translocation and release pathway of uracil/H+ by FurE, using path collective variable, funnel metadynamics and rationally designed mutational analysis. Our study reveals a step-wise, induced-fit, mechanism of ordered sequential transport of proton and uracil, which in turn suggests that the FurE symporter, and probably structurally similar transporters, functions as a multi-step gated pore, rather than employing ‘rocking’ of compact domains, as generally proposed for APC transporters. In addition, our work further supports the emerging concept that specific elements of cytosolic terminal regions of transporters might be functionally important.

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