Glutenase enzyme assays
View Patent ↗Administering an effective dose of glutenase to a Celiac or dermatitis herpetiformis patient reduces levels of toxic gluten oligopeptides, thereby attenuating or eliminating the damaging effects of gluten.
1. A method of determining the therapeutic efficacy of a candidate glutenase enzyme, the method comprising:
detecting the ability of a candidate enzyme to digest one or more peptides selected from the group consisting of (SEQ ID NO:4) FLQPQQPFPQQPQQPYPQQPQQPFPQ; (SEQ ID NO:5) IQPQQPAQL; (SEQ ID NO:27) QQPQQPYPQ; and (SEQ ID NO:28) SPQQQFPQ to fragments that are non-toxic to Celiac sprue patients.
2. The method according to claim 1 , further comprising the step of detecting the ability of a candidate enzyme to digest the peptide (SEQ ID NO:12) LQLQPFPQPQLPYPQPQLPYPQPQLPYPQ PQPF.
3. The method of claim 1 , further comprising performing an HPLC-trace to determine the products of digestion.
4. The method according to claim 1 , wherein the detecting step comprises mixing said one or more peptides with said candidate enzyme in a reaction mixture and perfusing into an intact intestine in an animal model.
5. The method of claim 4 , further comprising performing an HPLC trace to determine the products of digestion.
6. The method of claim 1 , wherein the detecting step comprises mixing said one or more peptides with said candidate enzyme in a reaction mixture comprising one or more intestinal brush border enzymes.
7. The method of claim 1 , wherein the detecting step comprises mixing said candidate enzyme in a reaction mixture comprising the peptides, wherein the peptides have been pre-treated with pepsin under physiological conditions.
8. The method of claim 1 , wherein the detecting step comprises mixing said candidate enzyme in a reaction mixture comprising the peptides, wherein the peptides have been pre-treated with trypsin, chymotrypsin, elastase, and carboxypeptidase.
9. The method of claim 7 ,wherein said peptides, prior to said mixing step, are further pre-treated with trypsin, chymotrypsin, elastase and carboxypeptidase.
10. The method of claim 1 , wherein the peptide is (SEQ ID NO:4) FLQPQQPFPQQPQQPYPQQPQQPFPQ.
11. The method of claim 1 , wherein the peptide is (SEQ ID NO:5) IQPQQPAQL.
12. The method of claim 1 , wherein the peptide is (SEQ ID NO:27) QQPQQPYPQ.
13. The method of claim 1 , wherein the peptide is (SEQ ID NO:28) SPQQQFPQ.