AgsE-deficient strain

Patent metadata

Field	Value
Publication number	US-10035986-B2
Application number	US-201314408043-A
Country	US
Kind code	B2
Filing date	Jul 19, 2013
Priority date	Jul 19, 2012
Publication date	Jul 31, 2018
Grant date	Jul 31, 2018

Abstract

The present invention relates to a mutant microbial host cell which is deficient in the production of the AgsE protein or in the production of an homologous thereof if compared with a parent microbial host cell which has not been modified and measured under the same conditions. It has been surprisingly found that when the mutant microbial host cell according to the invention is used in a method to produce a compound of interest, for example an enzyme, an improved yield of said compound is obtained if compared to a method in which a parent host cell which has not been modified is used when measured under the same conditions.

First claim

The invention claimed is:

1. A mutant filamentous fungal host cell which has been modified in the genome thereof to result in a deficiency in production of a polypeptide selected from the group consisting of:
a. a polypeptide comprising the amino acid sequence of SEQ ID NO: 3 or a polypeptide at least 90% identical thereto and having an activity of the amino acid sequence of SEQ ID NO: 3;
b. a mature polypeptide comprising the amino acid sequence of SEQ ID NO: 4 or a polypeptide at least 90% identical thereto and having an activity of the mature polypeptide; and
c. a polypeptide encoded by the polynucleotide sequence of SEQ ID NO: 1 or 2 or encoded by a polynucleotide at least 90% identical to SEQ ID NO: 1 or 2, wherein said polypeptide encoded by a polynucleotide at least 90% identical to SEQ ID NO: 1 or 2 has an activity of the polypeptide encoded by the polynucleotide sequence of SEQ ID NO: 1 or 2;
compared with a parent filamentous fungal host cell which has not been modified and measured under the same conditions;
wherein the modification in the genome of the mutant filamentous fungal host cell is selected from:
i) a full or partial deletion of SEQ ID NO: 1 or a polynucleotide at least 90% identical to SEQ ID NO: 1;
ii) a full or partial replacement of SEQ ID NO: 1 or a polynucleotide at least 90% identical to SEQ ID NO: 1 with a polynucleotide sequence which does not code for a polypeptide as defined in a to c or which codes for a partially or fully inactive form of a polypeptide as defined in a to c; and
iii) a disruption of SEQ ID NO: 1 or a polynucleotide at least 90% identical to SEQ ID NO: 1 by the insertion of one or more nucleotides in the polynucleotide sequence, wherein the disruption results in partial or full inactivation of a polypeptide as defined in a to c;
wherein the mutant filamentous fungal host cell further comprises at least one recombinant polynucleotide coding for an enzyme of interest or at least one recombinant polynucleotide coding for a polypeptide involved in production of a metabolite of interest, wherein the enzyme of interest or the metabolite of interest is a compound which can be produced and isolated from the host cell on an industrial scale and
wherein the activity of the polypeptide defined by a to c is an enzymatic activity which is α-1,3-glucan synthase activity [EC 2.4.1.183].

2. The mutant filamentous fungal host cell according to claim 1 , wherein the modification results in:
a) a reduced or no production of a polypeptide defined by a to c compared to the parent filamentous fungal host cell that has not been modified, when analysed under the same conditions and/or
b) a polypeptide derived from the polypeptide as defined in a to c with decreased or no activity compared to the parent filamentous fungal host cell that has not been modified, when analysed under the same conditions.

3. The mutant filamentous fungal host cell according to claim 1 , wherein the mutant filamentous fungal host cell
a. produces less of a polypeptide defined by a to c or it produces no polypeptide defined by a to c compared with the parent filamentous fungal host cell which has not been modified and measured under the same conditions; and/or
b. produces a polypeptide derived from the polypeptide defined by a to c with decreased or no activity compared to the parent filamentous fungal host cell that has not been modified, when analysed under the same conditions.

4. The mutant filamentous fungal host cell according claim 1 , wherein the mutant filamentous fungal host cell produces at least 5% less polypeptide defined by a to c compared with the parent filamentous fungal host cell which has not been modified and measured under the same conditions.

5. The mutant filamentous fungal host cell according to claim 1 , wherein the mutant filamentous fungal host cell produces a polypeptide derived from the polypeptide defined by a to c with at least 5% less enzymatic activity, compared with the parent filamentous fungal host cell which has not been modified and measured under the same conditions.

6. The mutant filamentous fungal host cell according to claim 1 , wherein the modification which results in a reduced or no production of a polypeptide defined by a to c is due to a reduced or no production of the mRNA encoding said polypeptide.

7. The mutant filamentous fungal host cell according to claim 1 , wherein the at least one recombinant polynucleotide coding for the enzyme of interest or the at least one recombinant polynucleotide coding for a polypeptide involved in the production of a metabolite of interest is operably linked to a promoter.

8. The mutant filamentous fungal host cell according to claim 7 , wherein the promoter is a carbohydrate inducible promoter.

9. The mutant filamentous fungal host cell according to claim 1 , wherein the mutant filamentous fungal host cell is selected from Aspergillus, Acremonium, Myceliophthora, Thielavia, Chrysosporium, Penicillium, Talaromyces, Rasamsonia, Fusarium , and Trichoderma.

10. The mutant filamentous fungal host cell according to claim 1 , wherein the mutant filamentous fungal host cell produces a polypeptide derived from the polypeptide defined by a to c with at least 99.9% less enzymatic activity, compared with the parent filamentous fungal host cell which has not been modified and measured under the same conditions.

11. The mutant filamentous fungal host cell according to claim 7 , wherein the promoter is an inducible promoter.

12. The mutant filamentous fungal host cell according to claim 8 , wherein the carbohydrate inducible promoter is selected from the group consisting of: a starch inducible promoter, a glucoamylase promoter, an acid stable amylase promoter, an alpha-amylase promoter, and a TAKA amylase promoter.

13. The mutant filamentous fungal host cell according claim 1 , wherein the mutant filamentous fungal host cell produces at least 99.9% less polypeptide defined by a to c compared with the parent filamentous fungal host cell which has not been modified and measured under the same conditions.

14. The mutant filamentous fungal host cell according to claim 1 , wherein the mutant filamentous fungal host cell is selected from Aspergillus niger, Aspergillus awamori, Aspergillus foetidus, Aspergillus sojae, Aspergillus fumigatus, Aspergillus oryzae, Acremonium alabamense, Myceliophthora thermophila, Thielavia terrestris, Chrysosporium lucknowense, Fusarium oxysporum, Rasamsonia emersonii, Talaromyces emersonii, Trichoderma reesei , and Penicillium chrysogenum.

Assignees

• Dsm Ip Assets Bv

Inventors

• VAN PEIJ NOëL NICOLAAS MARIA ELISABETH
• Beishuizen Martina
• Van De Vondervoort Peter Jozef Ida

Classifications

• C12Y301/01004

  Phospholipase A2 (3.1.1.4) · CPC title

• C12Y301/01003

  Triacylglycerol lipase (3.1.1.3) · CPC title

• C12Y204/01183

  Alpha-1,3-glucan synthase (2.4.1.183) · CPC title

• C12Y101/03004

  Glucose oxidase (1.1.3.4) · CPC title

• C12P21/02 Primary

  having a known sequence of two or more amino acids, e.g. glutathione · CPC title