Enzymatic modification of phospholipids in food
A phospholipase A1 characterized by having an sn1/sn2 specificity ratio of about 55/45 or greater wherein said phospholipase A1 has a lysophospholipase/phospholipase activity ratio of less than 0.02 is presented in conjunction with methods for use in lipid containing food matrix, baking and making dough with the phospholipase and also including baking improvers using the disclosed phospholipase A1.
1. A method of making a dough, said method comprising admixing a dough component selected from the group consisting of flour, salt, water, sugar, fat, lecithin, oil, emulsifier and yeast with an isolated polypeptide comprising a phospholipase A1 characterized by having an sn1/sn2 specificity ratio of about 55/45 or greater wherein said phospholipase A1 has a lysophospholipase/phospholipase activity ratio of less than 0.02 and/or a NALPE/NAPE activity ratio of less than 0.12.
2. The method of claim 1 wherein the sn1/sn2 specificity ratio is about 60/40, 70/30, 80/20, 90/10, 95/5 or 99/1.
3. The method of claim 2 wherein the sn1/sn2 specificity ratio is about 74/26 or 89/11.
4. The method of claim 3 wherein the lysophospholipase/phospholipase activity ratio is less than 0.009.
5. The method of claim 4 wherein the lysophospholipase/phospholipase activity ratio is less than 0.008.
6. The method of claim 5 wherein the lysophospholipase/phospholipase activity ratio is less than 0.007.
7. The method of claim 6 wherein the lysophospholipase/phospholipase activity ratio is less than 0.006.
8. The method of claim 7 wherein the lysophospholipase/phospholipase activity ratio is less than 0.005.
9. The method of claim 8 wherein the lysophospholipase/phospholipase activity ratio is less than 0.004.
10. The method of claim 9 wherein the lysophospholipase/phospholipase activity ratio is less than 0.003.
11. The method of claim 10 wherein the lysophospholipase/phospholipase activity ratio is less than 0.002.
12. The method of claim 11 wherein the lysophospholipase/phospholipase activity ratio is less than 0.001.
13. The method of claim 12 wherein the sn1/sn2 specificity ratio is about 60/40, 70/30, 80/20, 90/10, 95/5 or 99/1.
14. The method of claim 13 wherein the sn1/sn2 specificity ratio is about 74/26 or 89/11.
15. The method of claim 14 wherein said phospholipase A1 is an enzyme comprising a protein sequence having at least 80% sequence identity to SEQ ID NO: 2, SEQ ID NO: 4, SEQ ID NO: 6, SEQ ID NO: 8, SEQ ID NO: 10 SEQ ID NO: 12, SEQ ID NO: 14 or SEQ ID NO: 16.
16. The method of claim 15 wherein said phospholipase A1 is an enzyme comprising a protein sequence having at least 80% sequence identity to SEQ ID NO: 6.
17. The method of claim 15 wherein said phospholipase A1 is an enzyme comprising a protein sequence having at least 90% sequence identity to SEQ ID NO: 2, SEQ ID NO: 4, SEQ ID NO: 6, SEQ ID NO: 8, SEQ ID NO: 10 SEQ ID NO: 12, SEQ ID NO: 14 or SEQ ID NO: 16.
18. The method of claim 17 wherein said phospholipase A1 is an enzyme comprising a protein sequence having at least 90% sequence identity to SEQ ID NO: 6.
19. The method of claim 15 wherein said phospholipase A1 is an enzyme comprising a protein sequence having at least 95% sequence identity to SEQ ID NO: 2, SEQ ID NO: 4, SEQ ID NO: 6, SEQ ID NO: 8, SEQ ID NO: 10 SEQ ID NO: 12, SEQ ID NO: 14 or SEQ ID NO: 16.
20. The method of claim 15 wherein said phospholipase A1 is an enzyme comprising a protein sequence having at least 95% sequence identity to SEQ ID NO: 6.
21. The method of claim 15 wherein said phospholipase A1 is an enzyme comprising a protein sequence having 100% sequence identity to SEQ ID NO: 2, SEQ ID NO: 4, SEQ ID NO: 6, SEQ ID NO: 8, SEQ ID NO: 10 SEQ ID NO: 12, SEQ ID NO: 14 or SEQ ID NO: 16.
22. The method of claim 21 wherein said phospholipase A1 is an enzyme comprising a protein sequence having 100% sequence identity to SEQ ID NO: 6.
23. The method of claim 1 further comprising adding at least one additional enzyme useful for improving dough and/or a baked product made therefrom.
24. The method of claim 23 wherein the additional enzyme is selected from the group consisting of amylase, cyclodextrin glucanotransferase, peptidase, transglutaminase, lipase, galactolipase, phospholipase which is different than said phospholipase A1, cellulase, hemicellulase, protease, protein disulfide isomerase, glycosyltransferase, peroxidase, lipoxygenase, laccase, and oxidase.
25. The method of claim 24 wherein said amylase is an exoamylase.
26. The method of claim 25 wherein said exoamylase is a maltogenic amylase.
27. The method of claim 26 wherein said exoamylase is a non-maltogenic amylase.
28. The method of claim 27 wherein said non-maltogenic amylase hydrolyses starch by cleaving off one or more linear malto-oligosaccharides, predominantly comprising from four to eight D-glucopyranosyl units, from the non-reducing ends of the side chains of amylopectin.
29. The method of claim 24 wherein said additional enzyme is a phospholipase.
30. The method of claim 29 wherein said phospholipase has galactolipase activity.
31. The method of claim 30 wherein said phospholipase comprises a protein according to SEQ ID NO: 17 and/or SEQ ID NO: 18.
32. The method of claim 1 further comprising adding an emulsifier.
33. The method of claim 1 wherein the emulsifier is selected from the group consisting of i) phospholipid emulsifiers such as selected from the group consisting of lecithin and lyso-lecithin; and ii) a non-phospholipid emulsifier selected from the group consisting of DATEM, a monoglyceride and a diglyceride.
34. The method of claim 1 wherein the phospholipase A1 is an enzyme comprising a protein sequence having at least 80% sequence identity to SEQ ID NO:5 or a phospholipase active fragment thereof.
35. The method of claim 34 wherein the active fragment is a mature form of SEQ ID NO:5.