Indolmycin resistance of Streptomyces coelicolor A3(2) by induced expression of one of its two tryptophanyl-tRNA synthetases.
Aminoacyl-tRNA synthetases, a family of enzymes essential for protein synthesis, are promising targets of antimicrobials. Indolmycin, a secondary metabolite of Streptomyces griseus and a selective inhibitor of prokaryotic tryptophanyl-tRNA synthetase (TrpRS), was used to explore the mechanism of inhibition and to explain the resistance of a naturally occurring strain. Streptomyces coelicolor A3(2), an indolmycin-resistant strain, contains two trpS genes encoding distinct TrpRS enzymes. We show that TrpRS1 is indolmycin-resistant in vitro and in vivo, whereas TrpRS2 is sensitive. The lysine (position 9) in the enzyme tryptophan binding site is essential for making TrpRS1 indolmycin-resistant. Replacement of lysine 9 by glutamine, which at this position is conserved in most bacterial TrpRS proteins, abolished the ability of the mutant trpS gene to confer indolmycin resistance in vivo. Molecular modeling suggests that lysine 9 sterically hinders indolmycin binding to the enzyme. Tryptophan recognition (assessed by k(cat)/K(M)) by TrpRS1 is 4-fold lower than that of TrpRS2. Examination of the mRNA for the two enzymes revealed that only TrpRS2 mRNA is constitutively expressed, whereas mRNA for the indolmycin-resistant TrpRS1 enzyme is induced when the cells are exposed to indolmycin.
Kitabatake
Makoto
M
RIKEN Genomic Sciences Center, 1-7-22 Suehiro-cho, Tsurumi, Yokohama 230-0045, Japan.
Ali
Kamilah
K
Demain
Arnold
A
Sakamoto
Kensaku
K
Yokoyama
Shigeyuki
S
Söll
Dieter
D
D000595:Amino Acid Sequence; D000900:Anti-Bacterial Agents / Q000494:pharmacology*; D003001:Cloning, Molecular; D024881:Drug Resistance, Bacterial; D007211:Indoles / Q000494:pharmacology*; D007527:Isoenzymes / Q000032:analysis / Q000235:genetics; D008961:Models, Structural; D008969:Molecular Sequence Data; D012333:RNA, Messenger / Q000032:analysis; D016415:Sequence Alignment; D013302:Streptomyces / Q000187:drug effects*; D014369:Tryptophan-tRNA Ligase / Q000032:analysis / Q000737:chemistry* / Q000235:genetics
D000900:Anti-Bacterial Agents; D007211:Indoles; D007527:Isoenzymes; D012333:RNA, Messenger; D014369:Tryptophan-tRNA Ligase; C001447:indolmycin