Membrane topology of the L-rhamnose-H+ transport protein (RhaT) from enterobacteria

Abstract

The L-rhamnose-H+ symporter (RhaT) is a 344-amino acid integral membrane protein, found in many Enterobacteria, which couples the uptake of the sugar L-rhamnose with the inward movement of protons. Based on its hydropathy profile and the application of von Heijne's "positive inside" rule (von Heijne, G. (1992) J. Mol. Biol. 225, 487-494), a model of the L-rhamnose-H+ symport protein (RhaT) is proposed containing 10 transmembrane helices with the NH2 and COOH termini in the periplasm. This model was tested by the creation of random β-lactamase (Bla) fusions. The data from 33 unique, randomly generated, RhaT-Bla fusions and from 5 site-specific fusions supported the proposed topology between transmembrane helices 2-10. However, the localization of the putative first hydrophilic loop and the NH2 terminus was not possible because the β-lactamase fusions in this region were shown to be unreliable indicators of the topology of RhaT.