Determination of a transmembrane segment using cysteine-scanning mutants of transposon Tn10-encoded metal-tetracycline/H+ antiporter.
The metal-tetracycline/H+ antiporter is a bacterial plasma membrane protein belonging to the 12-transmembrane transporter family. The boundaries of membrane-embedded domains can be clearly determined by means of a method involving cysteine-scanning mutants and their reactivity to N-ethylmaleimide. The results for 37 cysteine-scanning mutants around putative transmembrane helix 9 indicate that the residues from 266 to 275 and from 296 to 298 form water extruding loops, while residues from 276 to 295 and from 299 to 302 are embedded in the membrane. This method is generally useful for determination of the transmembrane regions of a polytopic membrane protein.
Kimura
T
T
Department of Cell Membrane Biology, Osaka University, Japan.
Suzuki
M
M
Sawai
T
T
Yamaguchi
A
A
D000595:Amino Acid Sequence; D017920:Antiporters / Q000737:chemistry* / Q000235:genetics; D001426:Bacterial Proteins / Q000737:chemistry* / Q000235:genetics; D002462:Cell Membrane / Q000378:metabolism; D003545:Cysteine / Q000235:genetics*; D004591:Electrophoresis, Polyacrylamide Gel; D004926:Escherichia coli / Q000378:metabolism; D005033:Ethylmaleimide / Q000378:metabolism; D008565:Membrane Proteins / Q000737:chemistry; D008958:Models, Molecular; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed / Q000235:genetics; D009154:Mutation; D011485:Protein Binding; D013753:Tetracycline Resistance / Q000235:genetics
D017920:Antiporters; D001426:Bacterial Proteins; D008565:Membrane Proteins; C079051:tetA protein, Bacteria; D003545:Cysteine; D005033:Ethylmaleimide