Glycosylation as a stabilizer for phytase

Patent metadata

Field	Value
Publication number	US-9894917-B2
Application number	US-201314377134-A
Country	US
Kind code	B2
Filing date	Feb 1, 2013
Priority date	Feb 7, 2012
Publication date	Feb 20, 2018
Grant date	Feb 20, 2018

Abstract

The present teachings provide modified enzymes, preferably phytases, which have increased stability, hypothesized to arise from increased glycosylation. The enzymes can be modified to introduce or increase the number of glycosylation sites in the amino acid sequence, or glycosylation can be increased by the use of specific host production methods, or both. The enzymes of the present teachings have an increased stability after treatment at elevated temperature, which can be measured by inactivity reversibility or percent recovery following a treatment such as heating. The enzymes of the present teachings find application for example in food, feed, and feed pelleting.

First claim

What is claimed is:

1. A phytase polypeptide comprising SEQ ID NO: 1 and variants at least 90% identical thereto having increased glycosylation when compared to a control phytase lacking increased glycosylation, and made in a deglycosylation-deficient filamentous fungus, wherein said phytase has increased stability and pelleting stability when compared to a phytase made in a filamentous fungus that is not deglycosylation-deficient.

2. The phytase polypeptide of claim 1 comprising one or more of the following substitutions: E121T, P394N, D386N, K202N and N204T, Q151N and P153S, P373T, Q76N with reference to the position numbering of SEQ ID NO:1.

3. The phytase polypeptide of claim 2 wherein said polypeptide has the amino acid sequence of SEQ NOs 5, 6, 7, 8, 9, 10 or 11.

4. The phytase polypeptide of claims 1 or 3 having 1 or more added glycosylation sites.

5. The phytase polypeptide of claim 4 wherein the stability comprises increased inactivity reversibility following exposure to an elevated temperature as compared to a control phytase lacking increased glycosylation, wherein the elevated temperature is at least 80° C.

6. The phytase of claim 5 wherein the inactivity reversibility is at least 1% higher than a control phytase lacking increased glycosylation.

7. The phytase of claim 6 wherein the inactivity reversibility occurs after processing into a food or feed pellet.

8. The phytase polypeptide of claim 7 wherein the stability comprises increased recovered activity following exposure to an elevated temperature of at least 80° C. as compared to a control phytase lacking the increased glycosylation.

9. The phytase of claim 8 wherein the increased recovered activity as compared to a control phytase lacking increased glycosylation is at least 20%.

10. The phytase polypeptide of claim 7 wherein the stability comprises recovered activity following exposure to an elevated temperature of at least 80° C., as compared to the phytase prior to the exposure at the elevated temperature.

11. The phytase of claim 10 wherein the recovered activity is at least 40%, as compared to the phytase prior to the exposure at the elevated temperature.

12. The phytase polypeptide according to claim 11 wherein the increased recovered activity occurs after processing into a food or feed pellet.

13. The phytase polypeptide according to claim 1 , wherein the filamentous fungus is an Aspergillus spp., a Fusarium spp., a Myceliophthora spp., or a Trichoderma spp.

14. The phytase polypeptide according to claim 13 , wherein the Aspergillus is A. niger, A. oryzae, A. nidulans, A. tubingensis , or A. awamori.

15. The phytase polypeptide according to claim 13 , wherein the Trichoderma is T. reesei.

16. The phytase polypeptide according to claim 1 wherein the phytase is contained in a granule, optionally wherein the granule is a multi-layered granule, optionally wherein the granule is contained in a pellet, and optionally wherein the pellet is contained in an animal feed.

Assignees

• Danisco Us Inc
• Danisco Us Inc

Inventors

• Gebert Mark S
• Lee Sang Kyu
• Johnson Mariliz
• Ward Michael

Classifications

• A23K20/189 Primary

  Enzymes · CPC title

• A23K20/147

  Polymeric derivatives, e.g. peptides or proteins · CPC title

• C12Y301/03

  Phosphoric monoester hydrolases (3.1.3) · CPC title

• A23K1/1653 Primary

  Human Necessities · mapped topic

• C12Y301/03008

  3-Phytase (3.1.3.8) · CPC title