The periplasmic-binding proteins in ATP-binding cassette systems (ABC Transporters) are responsible for the capture and delivery of ligands to their specific transporters, triggering a series of ATP-driven conformational changes that leads to the transport of the ligand. Structurally consisting of two lobes, the proteins change conformation after interaction with the ligand. The structure of the molybdate-binding protein (ModA) from Xanthomonas citri, bound to molybdate, was previously solved by our group and an interdomain interaction, mediated by a salt bridge between K127 and D59, apparently supports the binding properties and keeps the domains closed. To determinate the importance of this interaction, we built two ModA mutants, K127S and D59A, and analysed their functional and structural properties. Based on a set of spectroscopic experiments, crystallisation trials, structure determination and molecular dynamics (MD) simulations, we showed that the salt bridge is essential to maintain the structure and binding properties. Additionally, the MD simulations revealed that this mutant adopted a more compact structure that packed down the ligand-binding pocket. From the closed bound to open structure, the positioning of the helices forming the dipole and the salt bridge are essential to induce an intermediate state.
Bibliographical noteFunding Information:
This work was supported by Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP) , Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq) and Associação Brasileira de Tecnologia de Luz Síncrotron (ABTLuS) . We also thankfully acknowledge the technical support from LNBio. The funders had no role in the study design, data collection and analysis, decision to publish, or preparation of the manuscript.
Copyright 2014 Elsevier B.V., All rights reserved.
- ABC transporters
- Domain interaction
- Periplasmic-binding proteins
- Xanthomonas citri