A Mutation in Na+-NQR Uncouples Electron Flow from Na+ Trans location in the Presence of K+

作者：Shea Michael E; Mezic Katherine G; Juarez Oscar; Barquera Blanca^*

来源：Biochemistry, 2015, 54(2): 490-496.

DOI：10.1021/bi501266e

The sodium-pumping NADH:ubiquinone oxidoreductase (Na+-NQR) is a bacterial respiratory enzyme that obtains energy from the redox reaction between NADH and ubiquinone and uses this energy to create an electrochemical Na(+)gradient across the cell membrane. A number of acidic residues in transmembrane helices have been shown to be important for Na+ translocation. One of these, Asp-397 in the NqrB subunit, is a key residue for Na+ uptake and binding. In this study, we show that when this residue is replaced with asparagine, the enzyme acquires a new sensitivity to K+; in the mutant, K+ both activates the redox reaction and uncouples it from the ion translocation reaction. In the wild-type enzyme, Na+ (or Li+) accelerates turnover while K+ alone does not activate. In the NqrB-D397N mutant, K+ accelerates the same internal electron transfer step (2Fe-2S -> FMNC) that is accelerated by Na+. This is the same step that is inhibited in mutants in which Na+ uptake is blocked. NqrB-D397N is able to translocate Na+ and Li+, but when K+ is introduced, no ion translocation is observed, regardless of whether Na+ or Li+ is present. Thus, this mutant, when it turns over in the presence of K+, is the first, and currently the only, example of an uncoupled Na+-NQR. The fact the redox reaction and ion pumping become decoupled from each other only in the presence of K+ provides a switch that promises to be a useful experimental tool.

• 出版日期2015-1-20