A unique hydrophobic cluster near the active site contributes to differences in borrelidin inhibition among threonyl-tRNA synthetases.

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Ruan B, Bovee ML, Sacher M, Stathopoulos C, Poralla K, Francklyn CS, Soll D

A unique hydrophobic cluster near the active site contributes to differences in borrelidin inhibition among threonyl-tRNA synthetases.

J Biol Chem. 2005 Jan 7;280(1):571-7. Epub 2004 Oct 26.

PubMed ID

15507440 [ View in PubMed

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Abstract

Borrelidin, a compound with anti-microbial and anti-angiogenic properties, is a known inhibitor of bacterial and eukaryal threonyl-tRNA synthetase (ThrRS). The inhibition mechanism of borrelidin is not well understood. Archaea contain archaeal and bacterial genre ThrRS enzymes that can be distinguished by their sequence. We explored species-specific borrelidin inhibition of ThrRSs. The activity of ThrRS from Sulfolobus solfataricus and Halobacterium sp. NRC-1 was inhibited by borrelidin, whereas ThrRS enzymes from Methanocaldococcus jannaschii and Archaeoglobus fulgidus were not. In Escherichia coli ThrRS, borrelidin binding induced a conformational change, and threonine was not activated as shown by ATP-PP(i) exchange and a transient kinetic assay measuring intrinsic tryptophan fluorescence changes. These assays further showed that borrelidin is a noncompetitive tight binding inhibitor of E. coli ThrRS with respect to threonine and ATP. Genetic selection of borrelidin-resistant mutants showed that borrelidin binds to a hydrophobic region (Thr-307, His-309, Cys-334, Pro-335, Leu-489, Leu-493) proximal to the zinc ion at the active site of the E. coli ThrRS. Mutating residue Leu-489 --> Trp reduced the space of the hydrophobic cluster and resulted in a 1500-fold increase of the K(i) value from 4 nM to 6 microm. An alignment of ThrRS sequences showed that this cluster is conserved in most organisms except for some Archaea (e.g. M. jannaschii, A. fulgidus) and some pathogens (e.g. Helicobacter pylori). This study illustrates how one class of natural product inhibitors affects aminoacyl-tRNA synthetase function, providing potentially useful information for structure-based inhibitor design.

DrugBank Data that Cites this Article

Drug Targets

Drug	Target	Kind	Organism	Pharmacological Action	Actions
Threonine	Threonine--tRNA ligase, cytoplasmic	Protein	Humans	Unknown	Not Available	Details