Selective inhibition of bacterial tryptophanyl-tRNA synthetases by indolmycin is mechanism-based

Tishan L. Williams, Yuhui W. Yin, Charles W. Carter

Research output: Contribution to journalArticle

16 Scopus citations

Abstract

Indolmycin is a natural tryptophan analog that competes with tryptophan for binding to tryptophanyl-tRNA synthetase (TrpRS) enzymes. Bacterial and eukaryotic cytosolic TrpRSs have comparable affinities for tryptophan (Km ~ 2 μM), and yet only bacterial TrpRSs are inhibited by indolmycin. Despite the similarity between these ligands, Bacillus stearothermophilus (Bs)TrpRS preferentially binds indolmycin ~1500-fold more tightly than its tryptophan substrate. Kinetic characterization and crystallographic analysis of BsTrpRS allowed us to probe novel aspects of indolmycin inhibitory action. Previous work had revealed that long range coupling to residues within an allosteric region called the D1 switch of BsTrpRS positions the Mg2+ ion in a manner that allows it to assist in transition state stabilization. The Mg2+ ion in the inhibited complex forms significantly closer contacts with non-bridging oxygen atoms from each phosphate group of ATP and three water molecules than occur in the (presumably catalytically competent) pre-transition state (preTS) crystal structures. We propose that this altered coordination stabilizes a ground state Mg2+ · ATP configuration, accounting for the high affinity inhibition of BsTrpRS by indolmycin. Conversely, both the ATP configuration and Mg2+ coordination in the human cytosolic (Hc) TrpRS preTS structure differ greatly from the BsTrpRS preTS structure. The effect of these differences is that catalysis occurs via a different transition state stabilization mechanism in Hc TrpRS with a yetto- be determined role for Mg2+. Modeling indolmycin into the tryptophan binding site points to steric hindrance and an inability to retain the interactions used for tryptophan substrate recognition as causes for the 1000-fold weaker indolmycin affinity to Hc TrpRS.

Original languageEnglish (US)
Pages (from-to)255-265
Number of pages11
JournalJournal of Biological Chemistry
Volume291
Issue number1
DOIs
StatePublished - Jan 1 2016

ASJC Scopus subject areas

  • Biochemistry
  • Molecular Biology
  • Cell Biology

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