Identification of selected spectrum antibiotics
Inventors
Peterson, Steven E. • Yenne, Samuel P. • ELLIS, Joseph Christopher • GUENTHER, Richard H.
Assignees
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Publication Number
US-11427860-B2
Publication Date
2022-08-30
Expiration Date
Abstract
Methods are disclosed for identifying antibacterial compounds which inhibit propagation of selected spectrum bacteria, which bacteria use specific tRNA to code for Ala, Met, Ser, or Leu that other bacteria do not use. In one embodiment, the selected spectrum bacteria use GCA to code for Ala, whereas other bacteria use a different codon to code for alanine. The methods involve determining whether putative inhibitors promote or inhibit complex formation between the tRNA and a bacterial ribosome, or between the tRNA and an aminoacyl synthetase. Compounds which promote or inhibit complex formation can disrupt protein production, which bacteria need to propagate. The identified antibacterial compounds can selectively inhibit bacterial propagation. By limiting their effects to the selected spectrum bacteria, these compounds can treat or prevent specific bacterial infections without disrupting the normal bacterial flora, the patients' microbiome, or causing antibacterial resistance.
Core Innovation
The invention relates to a method of identifying, by screening, a specific inhibitor of RNA translation in selected spectrum bacteria using nucleic acid molecules that are portions of a tRNA anticodon stem loop. The screened nucleic acid molecule is a single stranded nucleic acid molecule no more than 20 nucleotides in length, provided with a label, and is associated with Gram negative bacteria that use GCA to code for alanine.
The nucleic acid molecule is contacted with a ribosome or an aminoacyl synthetase capable of binding to the nucleic acid molecule. The screening evaluates whether a test compound inhibits or promotes binding of the labeled nucleic acid molecule to the ribosome or the aminoacyl synthetase, and binding is detected through a change in the label from an unbound state to a bound state.
The disclosed approach provides selectivity by targeting an organism-specific tRNA anticodon stem-loop fragment that is shared among selected spectrum pathogens but not used by most other bacteria. The method identifies compounds that selectively inhibit RNA translation in bacteria that use GCA for Ala, but will not inhibit RNA translation in bacteria that use alternative codons for Ala.
Claims Coverage
The partial content provides one independent claim (clm-00001). Across the referenced dependent features, the inventive features cover a screening method using a short labeled tRNA anticodon stem-loop fragment, selection of Gram negative bacteria that use GCA for alanine, detection of binding changes to a ribosome or aminoacyl synthetase, and codon-specific selectivity for Ala translation inhibition.
Screening for an inhibitor of RNA translation in selected spectrum bacteria
A method of identifying, by screening, a specific inhibitor of RNA translation in selected spectrum bacteria by contacting a labeled nucleic acid molecule portion of a tRNA anticodon stem loop with a ribosome or an aminoacyl synthetase capable of binding, and detecting inhibition or promotion of binding of the nucleic acid molecule and the ribosome or aminoacyl synthetase by the test compound.
Gram negative selection requiring GCA to code for alanine
The selected spectrum bacteria is a Gram negative bacteria which uses GCA to code for alanine, and the inhibition or promotion of binding indicates whether the test compound is useful for inhibiting Gram negative bacterial propagation.
Short single stranded labeled tRNA anticodon stem-loop fragment
The nucleic acid molecule is a single stranded nucleic acid molecule no more than 20 nucleotides in length, comprising a portion of the tRNA anticodon stem loop associated with Gram negative bacteria that use GCA to code for alanine.
Detect label change between unbound and bound states
The label changes from an unbound state to a bound state when the nucleic acid molecule binds to the ribosome or aminoacyl synthetase, and the detection step identifies whether a change in the label occurs.
Codon-specific selectivity for Ala via GCA
The method identifies compounds that selectively inhibit RNA translation in bacteria that use GCA to encode for Ala, but will not inhibit RNA translation in bacteria that use alternative codons for Ala.
The claim coverage centers on a codon-selective screening approach that uses a short, labeled tRNA anticodon stem-loop fragment from Gram negative bacteria that use GCA for alanine, detects label changes reporting binding to a ribosome or aminoacyl synthetase in the presence of test compounds, and identifies compounds that selectively inhibit RNA translation in GCA-dependent Ala bacteria while not inhibiting bacteria using alternative alanine codons.
Stated Advantages
Identifies compounds that selectively inhibit RNA translation in bacteria that use GCA to encode for Ala.
Does not inhibit RNA translation in bacteria that use alternative codons for Ala.
Provides a basis for determining whether a test compound is useful for inhibiting Gram negative bacterial propagation.
Documented Applications
Screening and identification of specific inhibitors of RNA translation in selected spectrum Gram negative bacteria that use GCA to encode alanine.
Use of test compounds identified by the screening to inhibit Gram negative bacterial propagation.
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