期刊
JOURNAL OF BIOCHEMISTRY
卷 142, 期 5, 页码 621-625出版社
OXFORD UNIV PRESS
DOI: 10.1093/jb/mvm169
关键词
multidrug resistance; SMR; membrane-impermeable SH-reagent (DTNB, AMS, biotin-PE-maleimide); ethidium efflux; dual membrane topology; EbrAB
EmrE in Escherichia coli belongs to the small multidrug resistance (SMR) transporter family. It functions as a homo-dimer, but the orientation of the two monomers in the membrane (membrane topology) is under debate. We expressed various single-cysteine EmrE mutants in E. coli cells lacking a major efflux transporter. Efflux from cells expressing the P55C or T56C mutant was blocked by the external application of membrane-impermeable SH-reagents. This is difficult to explain by the parallel topology configuration, because Pro55 and Thr56 are considered to be located in the cytoplasm. From both the periplasm and the cytoplasm, biotin-PE-maleimide, a bulky membrane-impermeable SH-reagent, could access the cysteine residue at the 25th position in the presence of transport substrates and at the 108th position. These observations support the anti-parallel topology in the membrane.
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