ATP hydrolysis and nucleotide exit enhance maltose translocation in the MalFGK2E importer

Bárbara Abreu,Carlos Cruz,A Sofia F Oliveira,Cláudio M Soares

doi:10.1038/s41598-021-89556-y

Bárbara Abreu, Carlos Cruz + Show 2 more

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https://doi.org/10.1038/s41598-021-89556-y

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Abstract

ATP binding cassette (ABC) transporters employ ATP hydrolysis to harness substrate translocation across membranes. The Escherichia coli MalFGK2E maltose importer is an example of a type I ABC importer and a model system for this class of ABC transporters. The MalFGK2E importer is responsible for the intake of malto-oligossacharides in E.coli. Despite being extensively studied, little is known about the effect of ATP hydrolysis and nucleotide exit on substrate transport. In this work, we studied this phenomenon using extensive molecular dynamics simulations (MD) along with potential of mean force calculations of maltose transport across the pore, in the pre-hydrolysis, post-hydrolysis and nucleotide-free states. We concluded that ATP hydrolysis and nucleotide exit trigger conformational changes that result in the decrease of energetic barriers to maltose translocation towards the cytoplasm, with a concomitant increase of the energy barrier in the periplasmic side of the pore, contributing for the irreversibility of the process. We also identified key residues that aid in positioning and orientation of maltose, as well as a novel binding pocket for maltose in MalG. Additionally, ATP hydrolysis leads to conformations similar to the nucleotide-free state. This study shows the contribution of ATP hydrolysis and nucleotide exit in the transport cycle, shedding light on ABC type I importer mechanisms.

Highlights

ATP binding cassette (ABC) transporters employ ATP hydrolysis to harness substrate translocation across membranes
Other simulation studies versed the communication between nucleotide binding domains (NBDs) and T MDs19,20 and concluded that the coupling helices and the Q-loops are essential for transmitting the impact of ATP hydrolysis from the NBDs to the transmembrane domains
In this work we simulated the E. coli MalFGK2E importer in different states of the transport cycle: the pre-hydrolysis state with ATP bound to the NBDs, the post-hydrolysis state with ADP and phosphate bound and an Apo state

Summary

Introduction

ATP binding cassette (ABC) transporters employ ATP hydrolysis to harness substrate translocation across membranes. In addition to the NBDs, ABC transporters contain transmembrane domains (TMDs) that bind and translocate the substrates[2,3] In this way, these proteins can either act as importers or exporters, depending on directionality of the transport. The Escherichia coli MalFGK2E importer is a type I importer located in the inner membrane responsible for the transport of maltose and malto-oligossacharides from the periplasm It is a heterodimer, constituted by five subunits: two copies of MalK, which constitute the NBD dimer, MalF and MalG are part of the TMDs and the substrate-binding protein – M alE7. It was discovered that MalE binding decreases the energetic barrier for MalFGK2E complex closure, promoting the movement of the MalF and MalG helices towards the c enter[14] In this way, the presence of MalE stabilizes the pre-translocation state.

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