Abstract
Proteins of the Smr family are the smallest multidrug transporters, about 110 amino acids long, that extrude various drugs in exchange with protons, thereby rendering bacteria resistant to these compounds. One of these proteins, EmrE, is an Escherichia coli protein, which has been cloned based on its ability to confer resistance to ethidium and methyl viologen and which has been extensively characterized. More than 60 genes coding for Smr proteins have been identified in several bacteria based on amino acid sequence similarity to the emrE gene. In this work we have analyzed the sequence similarity among these homologues and identified some distinct signature sequence elements and several fully conserved residues. Five of these homologues, from human pathogens Mycobacterium tuberculosis, Bordetella pertussis, and Pseudomonas aeruginosa and from Escherichia coli, were cloned into an E. coli expression system. The proteins were further characterized and show varying degrees of methyl viologen uptake into proteoliposomes and [(3)H]TPP binding in solubilized membranes. The homologues can also form mixed oligomers with EmrE that exhibit intermediate binding characteristics. A comparative study of various homologous proteins provides a tool for deciphering structure-function relationship and monomer-monomer interaction in multidrug transporters and in membrane proteins in general.
Highlights
Cestor and are the outcome of the evolutionary process of natural selection. They provide an excellent pool of natural mutants to study structure-function relationships in multidrug transporters. Some of these homologues are found in human pathogens such as Mycobacterium tuberculosis and Pseudomonas aeruginosa, and may be clinically important when dealing with multidrug resistant organisms
Sequence Homology Analysis of the Smr Family—A simple BLAST [18] of EmrE against the available sequence data bases reveals over 60 sequences with high similarity to EmrE, all belonging to the Eubacteria kingdom
We analyzed the sequence similarities among 63 sequences that were found to posses basic motifs associating them with the Smr family
Summary
Proteins of the Smr family are the smallest multidrug transporters, about 110 amino acids long, that extrude various drugs in exchange with protons, thereby rendering bacteria resistant to these compounds. One of these proteins, EmrE, is an Escherichia coli protein, which has been cloned based on its ability to confer resistance to ethidium and methyl viologen and which has been extensively characterized. They provide an excellent pool of natural mutants to study structure-function relationships in multidrug transporters Some of these homologues are found in human pathogens such as Mycobacterium tuberculosis and Pseudomonas aeruginosa, and may be clinically important when dealing with multidrug resistant organisms. The approach developed provides tools for analysis of structure-function relationships and monomer-monomer interactions
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