The TolC protein of Escherichia coli serves as a cell-surface receptor for the newly characterized TLS bacteriophage.
The TolC protein of Escherichia coli is implicated in a variety of diverse cellular functions, including antibiotic efflux and alpha-hemolysin secretion. An incidental role of TolC is to facilitate the entry of the bacteriophage TLS and colicin E1 into the bacterial cell. Despite the resolution of TolC's atomic structure, the roles of specific residues in its diverse functions are unknown. Here, we describe a genetic strategy for isolating missense tolC mutations that abolish the bacteriophage receptor activity of the TolC protein without influencing its role in antibiotic efflux. These spontaneous mutations affected two regions of the TolC protein and included base-pair substitutions, insertions, and deletions. Comparison of the TolC sequence with those of its homologues revealed two hypervariable stretches that were predicted to represent loops. Interestingly, all but one of the TolC alterations preventing phage binding were located in these two hypervariable regions, which are likely to be exposed on the cell surface. This was substantiated by the recently solved three-dimensional structure of TolC. Curiously, all the phage-resistant TolC mutants showed varying degrees of resistance to colicin E1, suggesting the involvement of overlapping regions of TolC in colicin E1 import and phage binding. The phage used in this study, TLS, was earlier reported as a strain of U3. However, we show here that, unlike the previously reported lipopolysaccharide-specific U3 phage, this phage displays a distinctly different host range and discrete morphological features and, in addition to utilizing TolC as receptor, it requires the inner core of a lipopolysaccharide.
German
G J
GJ
Department of Microbiology, Arizona State University, Tempe, AZ 85287, USA.
Misra
R
R
D000327:Adsorption; D000595:Amino Acid Sequence; D001425:Bacterial Outer Membrane Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D001435:Bacteriophages / Q000378:metabolism* / Q000472:pathogenicity; D003087:Colicins / Q000378:metabolism; D018360:Crystallography, X-Ray; D004352:Drug Resistance, Microbial; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism* / Q000821:virology*; D029968:Escherichia coli Proteins; D017353:Gene Deletion; D005798:Genes, Bacterial / Q000235:genetics; D008070:Lipopolysaccharides / Q000378:metabolism; D026901:Membrane Transport Proteins; D008958:Models, Molecular; D008969:Molecular Sequence Data; D020125:Mutation, Missense / Q000235:genetics; D010641:Phenotype; D011487:Protein Conformation; D021381:Protein Transport; D011991:Receptors, Virus / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*
D001425:Bacterial Outer Membrane Proteins; D003087:Colicins; D029968:Escherichia coli Proteins; D008070:Lipopolysaccharides; D026901:Membrane Transport Proteins; D011991:Receptors, Virus; C073126:tolC protein, E coli