Roles of acidic residues in the hydrophilic loop regions of metal-tetracycline/H+ antiporter Tet(K) of Staphylococcus aureus.
Three transmembrane glutamic acid residues play essential roles in the metal-tetracycline/H+ antiporter Tet(K) of Staphylococcus aureus [Fujihira et al., FEBS Lett. 391 (1996) 243-246]. In the putative hydrophilic loop region of the Tet(K) and Tet(L) proteins, six acidic residues are conserved. Asp74, Asp200, Asp318 and Glu381 are located on the putative cytoplasmic side, and Asp39 and Glu345 on the putative periplasmic side. These residues were replaced by a neutral amino acid residue or a charge-conserved one. In contrast to the transmembrane glutamic acid residues, the replacement of the two glutamic acid residues (Glu345 and Glu381) did not affect the tetracycline resistance level. Out of the other four aspartic acid residues, the only essential residue is Asp318, any replacement of which resulted in complete loss of the tetracycline resistance and transport activity. Asp318 is located in cytoplasmic loop 10-11 in the putative 14-transmembrane-segment topology of Tet(K). In the case of the tetracycline exporters of Gram-negative bacteria, the only essential acidic residue in the cytoplasmic loop region is located in loop 2-3 [Yamaguchi et al., Biochemistry 31 (1992) 8344-8348]. It may be a general role for tetracycline efflux proteins that three transmembrane and one cytoplasmic acidic residues are mandatory for the tetracycline transport function.
Fujihira
E
E
Department of Cell Membrane Biology, Institute of Scientific and Industrial Research, Osaka University, Japan.
Kimura
T
T
Yamaguchi
A
A
D000595:Amino Acid Sequence; D017920:Antiporters / Q000235:genetics* / Q000378:metabolism; D001426:Bacterial Proteins / Q000235:genetics* / Q000378:metabolism; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D017421:Sequence Analysis; D013211:Staphylococcus aureus / Q000378:metabolism*; D013753:Tetracycline Resistance
D017920:Antiporters; D001426:Bacterial Proteins; C079051:tetA protein, Bacteria