High-level chromosomally mediated tetracycline resistance in Neisseria gonorrhoeae results from a point mutation in the rpsJ gene encoding ribosomal protein S10 in combination with the mtrR and penB resistance determinants.
Neisseria gonorrhoeae becomes resistant to tetracycline by two major mechanisms: expression of a plasmid-encoded TetM protein and mutations in endogenous genes (chromosomally mediated resistance). Early studies by Sparling and colleagues (P. F. Sparling F. A. J. Sarubbi, and E. Blackman, J. Bacteriol. 124:740-749, 1975) demonstrated that three genes were involved in high-level chromosomally mediated tetracycline resistance (MIC of tetracycline > or = 2 microg/ml): ery-2 (now referred to as mtrR), penB, and tet-2. While the identities of the first two genes are known, the tet-2 gene has not been identified. We cloned the tet-2 gene, which confers tetracycline resistance, from tetracycline-resistant clinical isolate N. gonorrhoeae FA6140 and show that resistance is due to a single point mutation (Val-57 to Met) in the rpsJ gene (rpsJ1) encoding ribosomal protein S10. Moreover, the identical mutation was found in six distinct tetracycline-resistant clinical isolates in which the MIC of tetracycline was > or =2 microg/ml. Site-saturation mutagenesis of the codon for Val-57 identified two other amino acids (Leu and Gln) that conferred identical levels of resistance as the Met-57 mutation. The mutation maps to the vertex of a loop in S10 that is near the aminoacyl-tRNA site in the structure of the 30S ribosomal subunit from Thermus thermophilus, and the residue equivalent to Val-57 in T. thermophilus S10, Lys-55, is within 8 to 9 A of bound tetracycline. These data suggest that large noncharged amino acids alter the rRNA structure near the tetracycline-binding site, leading to a lower affinity of the antibiotic.
Hu
Mei
M
Department of Pharmacology, University of North Carolina at Chapel Hill, 27599-7365, USA.
Nandi
Sobhan
S
Davies
Christopher
C
Nicholas
Robert A
RA
D000483:Alleles; D000595:Amino Acid Sequence; D005798:Genes, Bacterial; D008826:Microbial Sensitivity Tests; D008957:Models, Genetic; D008958:Models, Molecular; D008969:Molecular Sequence Data; D009344:Neisseria gonorrhoeae / Q000187:drug effects* / Q000235:genetics / Q000302:isolation & purification; D017354:Point Mutation; D012269:Ribosomal Proteins / Q000737:chemistry / Q000235:genetics*; D017386:Sequence Homology, Amino Acid; D013753:Tetracycline Resistance / Q000235:genetics*
D012269:Ribosomal Proteins; C029537:ribosomal protein S10