HarA polypeptides and nucleic acids, and related methods and uses thereof
OwnerJohn P. Mueller, Eric T. Baimainventors
Life of this patent
Estimating the term…
Abstract
The present invention relates to harA polypeptides and nucleic acids encoding harA polypeptides and mutants thereof, derived from Enterococcus faecalis and Bacillus subtilis, as well as methods of using the polypeptides and nucleic acids in screening and diagnostic methods.
Claims17
A method of screening a compound to determine whether the compound binds to a harA polypeptide, which comprises:
- contacting the harA polypeptide with the compound under conditions suitable for binding;
- and detecting whether the compound binds to the harA polypeptide, wherein the harA polypeptide comprises an amino acid sequence at least 75% identical to (a) the amino acid sequence shown in FIG. 2 (SEQ ID NO:
- 2) or (b) the amino acid sequence shown in FIG. 4 (SEQ ID NO:
- 4).
A method of screening a compound to determine whether the compound inhibits harA activity, which comprises:
- measuring the activity of a harA polypeptide (i) in the absence and (ii) in the presence of the compound, under conditions suitable for measuring harA activity;
- wherein a decrease in harA activity in the presence of the compound compared to the harA activity in the absence of the compound identifies the compound as inhibiting harA activity, wherein the harA polypeptide comprises an amino acid sequence at least 75% identical to (a) the amino acid sequence shown in FIG. 2 (SEQ ID NO:
- 2) or (b) the amino acid sequence shown in FIG. 4 (SEQ ID NO:
- 4).
The use of a hygromycin A-resistant strain of E. faecalis or B. subtilis to determine whether an antibacterial agent is effective in treating organisms which exhibit harA-mediated drug resistance.
The use of a nucleic acid encoding a harA polypeptide to identify an organism containing a harA gene, wherein the nucleic acid comprises a nucleotide sequence selected from the group consisting of:
- (a) a nucleotide sequence which is at least 75% identical to (i) the nucleotide sequence shown in FIG. 1 (SEQ ID NO:
- 1) or (ii) the nucleotide sequence shown in FIG. 3 (SEQ ID NO:
- 3);
- (b) a nucleotide sequence complementary to the nucleotide sequence of (a);
- and
- (c) a nucleotide sequence that is degenerate to the nucleotide sequence of (a) or (b).
A recombinant vector comprising a nucleic acid encoding a harA polypeptide, wherein the nucleic acid comprises a nucleotide sequence selected from the group consisting of:
- (a) a nucleotide sequence which is at least 75% identical to (i) the nucleotide sequence shown in FIG. 1 (SEQ ID NO:
- 1) or (ii) the nucleotide sequence shown in FIG. 3 (SEQ ID NO:
- 3);
- (b) a nucleotide sequence complementary to the nucleotide sequence of (a);
- and
- (c) a nucleotide sequence that is degenerate to the nucleotide sequence of (a) or (b).
A recombinant vector comprising nucleic acid encoding a harA polypeptide, wherein the nucleic acid encoding the harA polypeptide comprises a nucleotide sequence comprising a chloramphenicol resistance insertion or an erythromycin resistance insertion.
A recombinant harA polypeptide comprising an amino acid sequence which is at least 75% identical to (i) the amino acid sequence shown in FIG. 2 (SEQ ID NO:
- 2) or (ii) the amino acid sequence shown in FIG. 4 (SEQ ID NO:
- 4).
Dependencies are inferred from the claim text.