GerN, an endospore germination protein of Bacillus cereus, is an Na+/H+-K+antiporter

GerN, a Bacillus cereus spore germination protein, exhibits homology to a w idely distributed group of putative cation transporters or channel proteins . GerN complemented the Na+-sensitive phenotype of an Escherichia coli muta nt that is deficient in Na+/H+ antiport activity (strain KNabc). GerN also reduced the concentration of K+ required to support growth of an E. coli mu tant deficient in K+ uptake (strain TK2420). In a fluorescence-based assay of everted E. coli KNabc membrane vesicles, GerN exhibited robust Na+/H+ an tiport activity, with a K-m for Na+ estimated at 1.5 mM at pH 8.0 and 25 mM at pH 7.0. Li+, but not K+, served as a substrate. GerN-mediated Na+/H+ an tiport was further demonstrated in everted vesicles as energy-dependent acc umulation of Na-22(+). GerN also used K+ as a coupling ion without complete ly replacing H+, as indicated by partial inhibition by K+ of H+ uptake into right-side-out vesicles loaded with Na+. K+ translocation as part of the a ntiport was supported by the stimulatory effect of intravesicular K+ on Na- 22(+) uptake by everted vesicles and the dependence of GerN-mediated Rb-86( +) efflux on the presence of Na+ in trans. The inhibitory patterns of proto nophore and thiocyanate were most consistent with an electrogenic Na+/H+-K antiport. GerN-mediated Na+/H+-K+ antiport was much more rapid than GerN-m ediated Na+/H+ antiport.