Defining membrane spanning domains and crucial membrane-localized acidic amino acid residues for K+ transport of a Kup/HAK/KT-type Escherichia coli potassium transporter.

Potassium (K+)-uptake transport proteins present in prokaryote and eukaryote cells are categorized into two classes; Trk/Ktr/HKT, K+ channel, and Kdp belong to the same superfamily, whereas the remaining K+-uptake family, Kup/HAK/KT has no homology to the others, and neither its membrane topology nor crucial residues for K+ uptake have been identified. We examined the topology of Kup from Escherichia coli. Results from the reporter fusion and cysteine labeling assays support a model with 12 membrane-spanning domains. A model for proton-coupled K+ uptake mediated by Kup has been proposed. However, this study did not show any stimulation of Kup activity at low pH and any evidence of involvement of the three His in Kup-mediated K+ uptake. Moreover, replacement of all four cysteines of Kup with serine did not abolish K+ transport activity. To gain insight on crucial residues of Kup-mediated K+ uptake activity, we focused on acidic residues in the predicted external and transmembrane regions, and identified four residues in the membrane regions required for K+ uptake activity. This is different from no membrane-localized acidic residues essential for Trk/Ktr/HKTs, K+ channels and Kdp. Taken together, these results demonstrate that Kup belongs to a distinct type of K+ transport system. [ABSTRACT FROM AUTHOR]