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US 6,428,971 B1Granted
Teichoic acid enzymes and assays
AssigneePharmacia & Upjohn Company
InventorsDean L. Shinabarger, Steven M. Swaney, Sara E. Morin
Life of this patent
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Abstract
This invention discloses a novel substrate and assay for the TAP enzyme. In addition novel DNA, proteins and peptides from genes and proteins associated with bacterial teichoic acid biosynthetic pathways, specifically the rodC gene and proteins and variations thereof are disclosed.
Claims54
A process for determining teichoic acid polymerase activity in a sample, the method comprising:
- combining CDP-glycerol, water, a lipoteichoic acid substrate from a bacterium selected from the group consisting of Staphylococcus aureus, Enterococcus faecalis, and Bacillus subtilis, and the sample to form a mixture;
- incubating the mixture under conditions effective for a teichoic acid polymerase in the sample, to cause the incorporation of at least one glycerol-3-phosphate moiety into the lipoteichoic acid substrate to form a lipoteichoic acid product having at least one additional glycerol-3-phosphate moiety incorporated therein, wherein the teichoic acid polymerase is encoded by DNA having at least about 70% homology with SEQ ID NO:1;
- and
- determining the presence or absence of the lipoteichoic acid product.
1 more element
A process for determining the presence or absence of lipoteichoic acid from a bacterium selected from the group consisting of Staphylococcus aureus, Enterococcus faecalis, and Bacillus subtilis in a sample, the method comprising:
- combining CDP-glycerol, water, a teichoic acid polymerase encoded by DNA having at least about 70% homology with SEQ ID NO:1, and the sample to form a mixture;
- incubating the mixture under conditions effective for the teichoic acid polymerase to cause the incorporation of at least one glycerol-3-phosphate moiety into a lipoteichoic acid present in the sample to form a lipoteichoic acid product having at least one additional glycerol-3-phosphate moiety incorporated therein;
- and
- determining the presence or absence of the lipoteichoic acid product.
1 more element
A process for screening teichoic acid polymerase inhibitors, the method comprising:
- combining CDP-glycerol, water, a lipoteichoic acid substrate from a bacterium selected from the group consisting of Staphylococcus aureus, Enterococcus faecalis, and Bacillus subtilis, a teichoic acid polymerase encoded by DNA having at least about 70% homology with SEQ ID NO:1, and a teichoic acid polymerase inhibitor to form a mixture;
- incubating the mixture under conditions effective, when the teichoic acid polymerase inhibitor is not present, for the teichoic acid polymerase to cause the incorporation of at least one glycerol-3-phosphate moiety into the lipoteichoic acid substrate to form a lipoteichoic acid product having at least one additional glycerol-3-phosphate moiety incorporated therein;
- and
- determining the presence or absence of the lipoteichoic acid product produced when the teichoic acid polymerase inhibitor is present in the mixture relative to the lipoteichoic acid product produced when the teichoic acid polymerase inhibitor is not present in the mixture.
1 more element
A process for monitoring enzymatic reactions catalyzed by teichoic acid polymerase, the method comprising:
- combining CDP-glycerol, water, a lipoteichoic acid substrate from a bacterium selected from the group consisting of Staphylococcus aureus, Enterococcus faecalis, and Bacillus subtilis, and a teichoic acid polymerase encoded by DNA having at least about 70% homology with SEQ ID NO:1, to form a mixture;
- incubating the mixture under conditions effective for the teichoic acid polymerase to cause the incorporation of at least one glycerol-3-phosphate moiety into the lipoteichoic acid substrate to form a lipoteichoic acid product having at least one additional glycerol-3-phosphate moiety incorporated therein;
- and
- determining the presence or absence of the lipoteichoic acid product.
1 more element
A process for determining teichoic acid polymerase activity in a sample, the method comprising:
- combining CDP-glycerol, water, a lipoteichoic acid substrate from a bacterium selected from the group consisting of Staphylococcus aureus, Enterococcus faecalis, and Bacillus subtilis, and the sample to form a mixture;
- incubating the mixture under conditions effective for a teichoic acid polymerase in the sample to cause the incorporation of at least one glycerol-3-phosphate moiety into the lipoteichoic acid substrate to form a lipoteichoic acid product having at least one additional glycerol-3-phosphate moiety incorporated therein, wherein the teichoic acid polymerase has an amino acid sequence with at least about 70% homology with SEQ ID NO:2;
- and
- determining the presence or absence of the lipoteichoic acid product.
1 more element
A process for determining the presence or absence of lipoteichoic acid from a bacterium selected from the group consisting of Staphylococcus aureus, Enterococcus faecalis, and Bacillus subtilis in a sample, the method comprising:
- combining CDP-glycerol, water, a teichoic acid polymerase having an amino acid sequence with at least about 70% homology with SEQ ID NO:2, and the sample to form a mixture;
- incubating the mixture under conditions effective for the teichoic acid polymerase to cause the incorporation of at least one glycerol-3-phosphate moiety into a lipoteichoic acid present in the sample to form a lipoteichoic acid product having at least one additional glycerol-3-phosphate moiety incorporated therein;
- and
- determining the presence or absence of the lipoteichoic acid product.
1 more element
A process for screening teichoic acid polymerase inhibitors, the method comprising:
- combining CDP-glycerol, water, a lipoteichoic acid substrate from a bacterium selected from the group consisting of Staphylococcus aureus, Enterococcus faecalis, and Bacillus subtilis, a teichoic acid polymerase having an amino acid sequence with at least about 70% homology with SEQ ID NO:2, and a teichoic acid polymerase inhibitor to form a mixture;
- incubating the mixture under conditions effective, when the teichoic acid polymerase inhibitor is not present, for the teichoic acid polymerase to cause the incorporation of at least one glycerol-3-phosphate moiety into the lipoteichoic acid substrate to form a lipoteichoic acid product having at least one additional glycerol-3-phosphate moiety incorporated therein;
- and
- determining the presence or absence of the lipoteichoic acid product produced when the teichoic acid polymerase inhibitor is present in the mixture relative to the lipoteichoic acid product produced when the teichoic acid polymerase inhibitor is not present in the mixture.
1 more element
A process for monitoring enzymatic reactions catalyzed by teichoic acid polymerase, the method comprising:
- combining CDP-glycerol, water, a lipoteichoic acid substrate from a bacterium selected from the group consisting of Staphylococcus aureus, Enterococcus faecalis, and Bacillus subtilis, and a teichoic acid polymerase having an amino acid sequence with at least about 70% homology with SEQ ID NO:2 to form a mixture;
- incubating the mixture under conditions effective for the teichoic acid polymerase to cause the incorporation of at least one glycerol-3-phosphate moiety into the lipoteichoic acid substrate to form a lipoteichoic acid product having at least one additional glycerol-3-phosphate moiety incorporated therein;
- and
- determining the presence or absence of the lipoteichoic acid product.
1 more element
Dependencies are inferred from the claim text.