Inhibitors of autoinducer transporters
AssigneePrinceton University
InventorsMichiko E. Taga, Bonnie L. Bassler, Douglas T. McKenzie
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Abstract
The present invention relates to the discovery of the lsr operon, the genes therein, and the polypeptides encoded by these genes. The present invention also includes strains with altered expression levels of the polypeptides encoded by the genes and the lsr operon relative to wild type cells. In some embodiments, the strains express a transporter that transports an autoinducer into the cell at a level higher than that of wild type cells. The present invention also includes methods for identifying compounds that modulate the transport of the autoinducer into cells.
Claims73
An isolated bacterial strain having an increased expression level of a transporter that transports an autoinducer into the strain relative to a wildtype strain, wherein the autoinducer is not an acyl-homoserine lactone and can interact with the Vibrio harveyi LuxQ protein, thereby inducing expression of a Vibrio harveyi operon comprising the luxCDABE genes.
A method for identifying a compound that modulates the response to a first autoinducer that is not an acyl-homoserine lactone and that can interact with the Vibrio harveyi LuxQ protein, thereby inducing expression of a Vibrio harveyi operon comprising the luxCDABE genes, comprising:
- obtaining a cell having increased expression of a transporter that transports the autoinducer into the cell, wherein the cell produces a detectable signal in response to the first autoinducer;
- measuring the response of the cell to the first autoinducer in the presence and absence of a test compound;
- and
- comparing the responses to determine whether the test compound modulates the response to the first autoinducer.
A method for screening a candidate compound for the ability to bind to a transporter that transports an autoinducer into a cell, wherein the autoinducer is not an acyl-homoserine lactone and can interact with the Vibrio harveyi LuxQ protein thereby inducing expression of a Vibrio harveyi operon comprising the luxCDABE genes, comprising:
- contacting the transporter with the candidate compound;
- and
- determining whether the compound specifically binds to the transporter.
A method of screening a candidate compound for the ability to modulate the binding of an autoinducer to a transporter, wherein the autoinducer is not an acyl-homoserine lactone and can interact with the Vibrio harveyi LuxQ protein thereby inducing expression of a Vibrio harveyi operon comprising the luxCDABE genes, comprising:
- comparing the binding of the autoinducer to the transporter in the presence and absence of the candidate compound;
- and
- determining whether the the extent of binding of the autoinducer to the transporter in the presence of the compound increases or decreases relative to the extent of binding in the absence of the compound.
A method of screening a candidate compound for the ability to bind a polypeptide comprising:
- contacting a polypeptide comprising a sequence selected from the group consisting of SEQ ID NOs. 37-40 and a sequence having at least 25% identity as determined through use of FASTA version 3.0t78 with the default parameters to one of the sequences of SEQ ID NOs.:
- 37-40 with the compound;
- and
- determining whether the compound specifically binds to the polypeptide.
An isolated bacterial strain comprising a mutation that inhibits the transport of an autoinducer that is not an acyl-homoserine lactone and that can interact with the Vibrio harveyi LuxQ protein thereby inducing expression of a Vibrio harveyi operon comprising the luxCDABE genes.
A bacterial strain that overexpresses or underexpresses a polypeptide comprising a sequence selected from the group consisting of SEQ ID NOs. 36-43 and a sequence having at least 25% identity to one of SEQ ID NOs:
- 36-43 relative to a wildtype strain.
A bacterial strain that overexpresses or underexpresses a polypeptide comprising a sequence selected from the group consisting of SEQ ID NOs. 36-43 and a sequence having at least 25% identity to one of SEQ ID NOs:
- 36-43 relative to a wildtype strain.
An isolated or purified nucleic acid comprising a sequence selected from the group consisting of SEQ ID NOs.:
- 28-35 and a fragment comprising at least 20 consecutive nucleotides of one of of SEQ ID NOs.:
- 28-35.
An isolated or purified nucleic acid comprising a fragment of one of SEQ ID NOs.:
- 28-35 that encodes a polypeptide that can facilitate the transport of an autoinducer into a cell, wherein the autoinducer is not an acyl-homoserine lactone and can interact with the Vibrio harveyi LuxQ protein thereby inducing expression of a Vibrio harveyi operon comprising the luxCDABE genes.
A recombinant vector comprising a sequence selected from the group consisting of SEQ ID NOs:
- 28-35 operably linked to a heterologous promoter.
An isolated or purified protein comprising a sequence selected from the group consisting of SEQ ID NOs.:
- 36-43 and a fragment comprising at least 10 consecutive amino acids of one of of SEQ ID NOs. 36-43.
An isolated or purified polypeptide comprising a fragment of one of SEQ ID NOs.:
- 36-43 that encodes a polypeptide that can facilitate the transport of an autoinducer into a cell, wherein the autoinducer is not an acyl-homoserine lactone and can interact with the Vibrio harveyi LuxQ protein thereby inducing expression of a Vibrio harveyi operon comprising the luxCDABE genes.
An antibody that binds to a polypeptide selected from the group consisting of SEQ ID NOs.:
- 36-43.
Dependencies are inferred from the claim text.