The Poly-Glutamate Motif of GmMATE4 Regulates Its Isoflavone Transport Activity.

Multidrug and toxic compound extrusion (MATE) transporters in eukaryotes have been characterized to be antiporters that mediate the transport of substrates in exchange for protons. In plants, alkaloids, phytohormones, ion chelators, and flavonoids have been reported to be the substrates of MATE transporters. Structural analyses have been conducted to dissect the functional significance of various motifs of MATE proteins. However, an understanding of the functions of the N- and C-termini has been inadequate. Here, by performing phylogenetic analyses and protein sequence alignment of 14 representative plant species, we identified a distinctive N-terminal poly-glutamate motif among a cluster of MATE proteins in soybean. Amongst them, GmMATE4 has the most consecutive glutamate residues at the N-terminus. A subcellular localization study showed that GmMATE4 was localized at the vacuolar membrane-like structure. Protein charge prediction showed that the mutation of the glutamate residues to alanine would reduce the negative charge at the N-terminus. Using yeast as the model, we showed that GmMATE4 mediated the transport of daidzein, genistein, glycitein, and glycitin. In addition, the glutamate-to-alanine mutation reduced the isoflavone transport capacity of GmMATE4. Altogether, we demonstrated GmMATE4 as an isoflavone transporter and the functional significance of the N-terminal poly-glutamate motif of GmMATE4 for regulating the isoflavone transport activity.