GENE PRODUCTS OF BACILLUS LICHENIFORMIS WHICH FORM ODOROUS SUBSTANCES AND IMPROVED BIOTECHNOLOGICAL PRODUCTION METHODS BASED THEREON
First Claim
1. A nucleic acid selected from the group consisting of:
- (a) nucleic acids coding for a gene product (putative branched-chain amino acid aminotransferase;
E.C. 2.6.1.42) involved in the synthesis of 2-methylbutyric acid and/or isobutyric acid and having a nucleotide sequence which shows at least 67% identity to the nucleotide sequence indicated in SEQ ID NO. 1;
(b) nucleic acids coding for a gene product (putative branched-chain amino acid aminotransferase;
E.C. 2.6.1.42) involved in the synthesis of 2-methylbutyric acid and/or isobutyric acid and having a nucleotide sequence which shows at least 78% identity to the nucleotide sequence indicated in SEQ ID NO. 3;
(c) nucleic acids speA coding for a gene product (lysine and/or arginine decarboxylase;
E.C. 4.1.1.18 or 4.1.1.19) involved in the synthesis of cadaverine and/or putrescine and having a nucleotide sequence which shows at least 78% identity to the nucleotide sequence indicated in SEQ ID No. 5;
(d) nucleic acids yugJ coding for a gene product (NADH-dependent butanol dehydrogenase A;
E.C. 1.1.1.-) involved in the synthesis of butanol and/or butyric acid and having a nucleotide sequence which shows at least 81% identity to the nucleotide sequence indicated in SEQ ID NO. 7;
(e) nucleic acids coding for a gene product (acyl-CoA dehydrogenase;
E.C. 1.3.99.-) involved in the synthesis of isovaleric acid, 2-methylbutyric acid, isobutyric acid, butanol and/or butyric acid and having a nucleotide sequence which shows at least 75% identity to the nucleotide sequence indicated in SEQ ID NO. 9;
(f) nucleic acids coding for a gene product (acyl-CoA dehydrogenase;
E.C. 1.3.99.-) involved in the synthesis of isovaleric acid, 2-methylbutyric acid, isobutyric acid, butanol and/or butyric acid and having a nucleotide sequence which shows at least 64% identity to the nucleotide sequence indicated in SEQ ID NO. 11;
(g) nucleic acids coding for a gene product (3-hydroxybutyryl-CoA dehydrogenase;
E.C. 1.1.1.157) involved in the synthesis of butanol and/or butyric acid and having a nucleotide sequence which shows at least 67% identity to the nucleotide sequence indicated in SEQ ID NO. 13;
(h) nucleic acids coding for a gene product (putative enoyl-Coa hydratase protein;
E.C. 4.2.1.17) involved in the synthesis of isovaleric acid, 2-methylbutyric acid and/or isobutyric acid and having a nucleotide sequence which shows at least 65% identity to the nucleotide sequence indicated in SEQ ID NO. 15;
(i) nucleic acids coding for a gene product (probable enoyl-(3-hydroxyisobutyryl)-coenzyme A hydrolase protein) involved in the synthesis of isovaleric acid, 2-methylbutyric acid and/or isobutyric acid and having a nucleotide sequence which shows at least 66% identity to the nucleotide sequence indicated in SEQ ID NO. 17;
(j) nucleic acids echA8 coding for a gene product (probable enoyl-CoA hydratase;
E.C. 4.2.1.17) involved in the synthesis of isovaleric acid, 2-methylbutyric acid and/or isobutyric acid and having a nucleotide sequence which shows at least 48% identity to the nucleotide sequence indicated in SEQ ID NO. 19;
(k) nucleic acids coding for a gene product (acyl-CoA dehydrogenase;
E.C. 1.3.99.-) involved in the synthesis of isovaleric acid, 2-methylbutyric acid and/or isobutyric acid and having a nucleotide sequence which shows at least 54% identity to the nucleotide sequence indicated in SEQ ID NO. 21;
(l) nucleic acids acsA coding for a gene product (acetyl-coenzyme A synthetase;
E.C. 6.2.1.1) involved in the synthesis of propionic acid and having a nucleotide sequence which shows at least 67% identity to the nucleotide sequence indicated in SEQ ID NO. 23;
(m) nucleic acids yngF coding for a gene product (3-hydroxybutyryl-CoA dehydratase;
E.C. 4.2.1.55) involved in the synthesis of butanol and/or butyric acid and having a nucleotide sequence which shows at least 68% identity to the nucleotide sequence indicated in SEQ ID NO. 25;
(n) nucleic acids yusJ coding for a gene product (acyl-CoA dehydrogenase;
E.C. 1.3.99.-) involved in the synthesis of isovaleric acid, 2-methylbutyric acid, isobutyric acid, butanol and/or butyric acid and having a nucleotide sequence which shows at least 77% identity to the nucleotide sequence indicated in SEQ ID NO. 27;
(o) nucleic acids ykwC coding for a gene product (hypothetical oxidoreductase;
E.C. 1.1.-.-) involved in the synthesis of 2-methylbutyric acid and/or isobutyric acid and having a nucleotide sequence which shows at least 77% identity to the nucleotide sequence indicated in SEQ ID NO. 29;
(p) nucleic acids coding for a gene product (probable phosphate butyryltransferase;
E.C. 2.3.1.19) involved in the synthesis of butanol and/or butyric acid and having a nucleotide sequence which shows at least 51% identity to the nucleotide sequence indicated in SEQ ID NO. 31;
(q) nucleic acids coding for a gene product (probable butyrate kinase;
E.C. 2.7.2.7) involved in the synthesis of butanol and/or butyric acid and having a nucleotide sequence which shows at least 77% identity to the nucleotide sequence indicated in SEQ ID NO. 33;
(r) nucleic acids acsA coding for a gene product (acetyl-coenzyme A synthetase;
E.C. 6.2.1.1) involved in the synthesis of propionic acid and having a nucleotide sequence which shows at least 79% identity to the nucleotide sequence indicated in SEQ ID NO. 35;
(s) nucleic acids ytcl coding for a gene product (acetate-CoA ligase;
E.C. 6.2.1.1) involved in the synthesis of propionic acid and having a nucleotide sequence which shows at least 74% identity to the nucleotide sequence indicated in SEQ ID NO. 37;
(t) nucleic acids speA coding for a gene product (lysine and/or arginine decarboxylase;
E.C. 4.1.1.18 or E.C. 4.1.1.19) involved in the synthesis of cadaverine and/or putrescine and having a nucleotide sequence which shows at least 68% identity to the nucleotide sequence indicated in SEQ ID NO. 39;
(u) nucleic acids ysiB coding for a gene product (probable Enoyl-CoA hydratase;
E.C. 4.2.1.17) involved in the synthesis of isovaleric acid, 2-methylbutyric acid and/or isobutyric acid and having a nucleotide sequence which shows at least 75% identity to the nucleotide sequence indicated in SEQ ID NO. 41;
(v) nucleic acids coding for a gene product (similar to 3-hydroxyacyl-CoA dehydrogenase;
E.C. 1.1.1.35) involved in the synthesis of 2-methylbutyric acid and having a nucleotide sequence which shows at least 76% identity to the nucleotide sequence indicated in SEQ ID NO. 43;
(w) nucleic acids coding for a gene product (2-oxoglutarate dehydrogenase E1 component;
E.C. 1.2.4.2) involved in the synthesis of isovaleric acid, 2-methylbutyric acid and/or isobutyric acid and having a nucleotide sequence which shows at least 80% identity to the nucleotide sequence indicated in SEQ ID NO. 45;
(x) nucleic acids yhfL coding for a gene product (probable acid-CoA ligase;
E.C. 6.2.1.-) involved in the synthesis of propionic acid and having a nucleotide sequence which shows at least 67% identity to the nucleotide sequence indicated in SEQ ID NO. 47; and
(y) nucleic acids ywhG coding for a gene product (agmatinase;
E.C. 3.5.1.11) involved in the synthesis of cadaverine and/or putrescine and having a nucleotide sequence which shows at least 85% identity to the nucleotide sequence indicated in SEQ ID NO. 49.
3 Assignments
0 Petitions
Accused Products
Abstract
The present invention relates to 25 hitherto undescribed genes of B. licheniformis and gene products derived therefrom and all sufficiently homologous nucleic acids and proteins thereof. They occur in five different metabolic pathways for the formation of odorous substances. The metabolic pathways in question are for the synthesis of: 1) isovalerian acid (as part of the catabolism of leucine), 2) 2-methylbutyric acid and/or isobutyric acid (as part of the catabolism of valine and/or isoleucine), 3) butanol and/or butyric acid (as part of the metabolism of butyric acid), 4) propyl acid (as part of the metabolism of propionate) and/or 5) cadaverine and/or putrescine (as parts of the catabolism of lysine and/or arginine). The identification of these genes allows biotechnological production methods to be developed that are improved to the extent that, to assist these nucleic acids, the formation of the odorous substances synthesized via these metabolic pathways can be reduced by deactivating the corresponding genes in the micro-organism used for the biotechnological production. In addition, these gene products are thus available for preparing reactions or for methods according to their respective biochemical properties.
86 Citations
30 Claims
-
1. A nucleic acid selected from the group consisting of:
-
(a) nucleic acids coding for a gene product (putative branched-chain amino acid aminotransferase;
E.C. 2.6.1.42) involved in the synthesis of 2-methylbutyric acid and/or isobutyric acid and having a nucleotide sequence which shows at least 67% identity to the nucleotide sequence indicated in SEQ ID NO. 1;
(b) nucleic acids coding for a gene product (putative branched-chain amino acid aminotransferase;
E.C. 2.6.1.42) involved in the synthesis of 2-methylbutyric acid and/or isobutyric acid and having a nucleotide sequence which shows at least 78% identity to the nucleotide sequence indicated in SEQ ID NO. 3;
(c) nucleic acids speA coding for a gene product (lysine and/or arginine decarboxylase;
E.C. 4.1.1.18 or 4.1.1.19) involved in the synthesis of cadaverine and/or putrescine and having a nucleotide sequence which shows at least 78% identity to the nucleotide sequence indicated in SEQ ID No. 5;
(d) nucleic acids yugJ coding for a gene product (NADH-dependent butanol dehydrogenase A;
E.C. 1.1.1.-) involved in the synthesis of butanol and/or butyric acid and having a nucleotide sequence which shows at least 81% identity to the nucleotide sequence indicated in SEQ ID NO. 7;
(e) nucleic acids coding for a gene product (acyl-CoA dehydrogenase;
E.C. 1.3.99.-) involved in the synthesis of isovaleric acid, 2-methylbutyric acid, isobutyric acid, butanol and/or butyric acid and having a nucleotide sequence which shows at least 75% identity to the nucleotide sequence indicated in SEQ ID NO. 9;
(f) nucleic acids coding for a gene product (acyl-CoA dehydrogenase;
E.C. 1.3.99.-) involved in the synthesis of isovaleric acid, 2-methylbutyric acid, isobutyric acid, butanol and/or butyric acid and having a nucleotide sequence which shows at least 64% identity to the nucleotide sequence indicated in SEQ ID NO. 11;
(g) nucleic acids coding for a gene product (3-hydroxybutyryl-CoA dehydrogenase;
E.C. 1.1.1.157) involved in the synthesis of butanol and/or butyric acid and having a nucleotide sequence which shows at least 67% identity to the nucleotide sequence indicated in SEQ ID NO. 13;
(h) nucleic acids coding for a gene product (putative enoyl-Coa hydratase protein;
E.C. 4.2.1.17) involved in the synthesis of isovaleric acid, 2-methylbutyric acid and/or isobutyric acid and having a nucleotide sequence which shows at least 65% identity to the nucleotide sequence indicated in SEQ ID NO. 15;
(i) nucleic acids coding for a gene product (probable enoyl-(3-hydroxyisobutyryl)-coenzyme A hydrolase protein) involved in the synthesis of isovaleric acid, 2-methylbutyric acid and/or isobutyric acid and having a nucleotide sequence which shows at least 66% identity to the nucleotide sequence indicated in SEQ ID NO. 17;
(j) nucleic acids echA8 coding for a gene product (probable enoyl-CoA hydratase;
E.C. 4.2.1.17) involved in the synthesis of isovaleric acid, 2-methylbutyric acid and/or isobutyric acid and having a nucleotide sequence which shows at least 48% identity to the nucleotide sequence indicated in SEQ ID NO. 19;
(k) nucleic acids coding for a gene product (acyl-CoA dehydrogenase;
E.C. 1.3.99.-) involved in the synthesis of isovaleric acid, 2-methylbutyric acid and/or isobutyric acid and having a nucleotide sequence which shows at least 54% identity to the nucleotide sequence indicated in SEQ ID NO. 21;
(l) nucleic acids acsA coding for a gene product (acetyl-coenzyme A synthetase;
E.C. 6.2.1.1) involved in the synthesis of propionic acid and having a nucleotide sequence which shows at least 67% identity to the nucleotide sequence indicated in SEQ ID NO. 23;
(m) nucleic acids yngF coding for a gene product (3-hydroxybutyryl-CoA dehydratase;
E.C. 4.2.1.55) involved in the synthesis of butanol and/or butyric acid and having a nucleotide sequence which shows at least 68% identity to the nucleotide sequence indicated in SEQ ID NO. 25;
(n) nucleic acids yusJ coding for a gene product (acyl-CoA dehydrogenase;
E.C. 1.3.99.-) involved in the synthesis of isovaleric acid, 2-methylbutyric acid, isobutyric acid, butanol and/or butyric acid and having a nucleotide sequence which shows at least 77% identity to the nucleotide sequence indicated in SEQ ID NO. 27;
(o) nucleic acids ykwC coding for a gene product (hypothetical oxidoreductase;
E.C. 1.1.-.-) involved in the synthesis of 2-methylbutyric acid and/or isobutyric acid and having a nucleotide sequence which shows at least 77% identity to the nucleotide sequence indicated in SEQ ID NO. 29;
(p) nucleic acids coding for a gene product (probable phosphate butyryltransferase;
E.C. 2.3.1.19) involved in the synthesis of butanol and/or butyric acid and having a nucleotide sequence which shows at least 51% identity to the nucleotide sequence indicated in SEQ ID NO. 31;
(q) nucleic acids coding for a gene product (probable butyrate kinase;
E.C. 2.7.2.7) involved in the synthesis of butanol and/or butyric acid and having a nucleotide sequence which shows at least 77% identity to the nucleotide sequence indicated in SEQ ID NO. 33;
(r) nucleic acids acsA coding for a gene product (acetyl-coenzyme A synthetase;
E.C. 6.2.1.1) involved in the synthesis of propionic acid and having a nucleotide sequence which shows at least 79% identity to the nucleotide sequence indicated in SEQ ID NO. 35;
(s) nucleic acids ytcl coding for a gene product (acetate-CoA ligase;
E.C. 6.2.1.1) involved in the synthesis of propionic acid and having a nucleotide sequence which shows at least 74% identity to the nucleotide sequence indicated in SEQ ID NO. 37;
(t) nucleic acids speA coding for a gene product (lysine and/or arginine decarboxylase;
E.C. 4.1.1.18 or E.C. 4.1.1.19) involved in the synthesis of cadaverine and/or putrescine and having a nucleotide sequence which shows at least 68% identity to the nucleotide sequence indicated in SEQ ID NO. 39;
(u) nucleic acids ysiB coding for a gene product (probable Enoyl-CoA hydratase;
E.C. 4.2.1.17) involved in the synthesis of isovaleric acid, 2-methylbutyric acid and/or isobutyric acid and having a nucleotide sequence which shows at least 75% identity to the nucleotide sequence indicated in SEQ ID NO. 41;
(v) nucleic acids coding for a gene product (similar to 3-hydroxyacyl-CoA dehydrogenase;
E.C. 1.1.1.35) involved in the synthesis of 2-methylbutyric acid and having a nucleotide sequence which shows at least 76% identity to the nucleotide sequence indicated in SEQ ID NO. 43;
(w) nucleic acids coding for a gene product (2-oxoglutarate dehydrogenase E1 component;
E.C. 1.2.4.2) involved in the synthesis of isovaleric acid, 2-methylbutyric acid and/or isobutyric acid and having a nucleotide sequence which shows at least 80% identity to the nucleotide sequence indicated in SEQ ID NO. 45;
(x) nucleic acids yhfL coding for a gene product (probable acid-CoA ligase;
E.C. 6.2.1.-) involved in the synthesis of propionic acid and having a nucleotide sequence which shows at least 67% identity to the nucleotide sequence indicated in SEQ ID NO. 47; and
(y) nucleic acids ywhG coding for a gene product (agmatinase;
E.C. 3.5.1.11) involved in the synthesis of cadaverine and/or putrescine and having a nucleotide sequence which shows at least 85% identity to the nucleotide sequence indicated in SEQ ID NO. 49. - View Dependent Claims (3)
-
-
2. A gene product selected from the group consisting of:
-
(a) gene products (putative branched-chain amino acid aminotransferase;
E.C. 2.6.1.42) involved in the synthesis of 2-methylbutyric acid and/or isobutyric acid and having an amino acid sequence which shows at least 73% identity to the amino acid sequence indicated in SEQ ID NO. 2;
(b) gene products (putative branched-chain amino acid aminotransferase;
E.C. 2.6.1.42) involved in the synthesis of 2-methylbutyric acid and/or isobutyric acid and having an amino acid sequence which shows at least 83% identity to the amino acid sequence indicated in SEQ ID NO. 4;
(c) gene products SpeA (lysine and/or arginine decarboxylase;
E.C. 4.1.1.18 or E.C. 4.1.1.19) involved in the synthesis of cadaverine and/or putrescine and having an amino acid sequence which shows at least 89% identity to the amino acid sequence indicated in SEQ ID NO. 6;
(d) gene products YugJ (NADH-dependent butanol dehydrogenase A;
E.C. 1.1.1.-) involved in the synthesis of butanol and/or butyric acid and having an amino acid sequence which shows at least 93% identity to the amino acid sequence indicated in SEQ ID NO. 8;
(e) gene products (acyl-CoA dehydrogenase;
E.C. 1.3.99.-) involved in the synthesis of isovaleric acid, 2-methylbutyric acid, isobutyric acid, butanol and/or butyric acid and having an amino acid sequence which shows at least 86% identity to the amino acid sequence indicated in SEQ ID NO. 10;
(f) gene products (acyl-CoA dehydrogenase;
E.C. 1.3.99.-) involved in the synthesis of isovaleric acid, 2-methylbutyric acid, isobutyric acid, butanol and/or butyric acid and having an amino acid sequence which shows at least 67% identity to the amino acid sequence indicated in SEQ ID NO. 12;
(g) gene products (3-hydroxybutyryl-CoA dehydrogenase;
E.C. 1.1.1.157) involved in the synthesis of butanol and/or butyric acid and having an amino acid sequence which shows at least 69% identity to the amino acid sequence indicated in SEQ ID NO. 14;
(h) gene products (putative enoyl-Coa hydratase protein;
E.C. 4.2.1.17) involved in the synthesis of isovaleric acid, 2-methylbutyric acid and/or isobutyric acid and having an amino acid sequence which shows at least 62% identity to the amino acid sequence indicated in SEQ ID NO. 16;
(i) gene products (probable enoyl-(3-hydroxyisobutytyl)-coenzyme A hydrolase protein) involved in the synthesis of isovaleric acid, 2-methylbutyric acid and/or isobutyric acid and having an amino acid sequence which shows at least 66% identity to the amino acid sequence indicated in SEQ ID NO. 18;
(j) gene products EchA8 (probable enoyl-CoA hydratase;
E.C. 4.2.1.17) involved in the synthesis of isovaleric acid, 2-methylbutyric acid and/or isobutyric acid and having an amino acid sequence which shows at least 52% identity to the amino acid sequence indicated in SEQ ID NO. 20;
(k) gene products (acyl-CoA dehydrogenase) involved in the synthesis of isovaleric acid, 2-methylbutyric acid and/or isobutyric acid and having an amino acid sequence which shows at least 65% identity to the amino acid sequence indicated in SEQ ID NO. 22;
(l) gene products AcsA (acetyl-coenzyme A synthetase;
E.C. 6.2.1.1) involved in the synthesis of propionic acid and having an amino acid sequence which shows at least 65% to the amino acid sequence indicated in SEQ ID NO. 24;
(m) gene products YngF (3-hydroxybutyryl-CoA dehydratase;
E.C. 4.2.1.55) involved in the synthesis of butanol and/or butyric acid and having an amino acid sequence which shows at least 69% identity to the amino acid sequence indicated in SEQ ID NO. 26;
(n) gene products YusJ (acyl-CoA dehydrogenase;
E.C. 1.3.99.-) involved in the synthesis of isovaleric acid, 2-methylbutyric acid, isobutyric acid, butanol and/or butyric acid and having an amino acid sequence which shows at least 86% identity to the amino acid sequence indicated in SEQ ID NO. 28;
(o) gene products YkwC (hypothetical oxidoreductase;
E.C. 1.1.-.-) involved in the synthesis of 2-methylbutyric acid and/or isobutyric acid and having an amino acid sequence which shows at least 85% identity to the amino acid sequence indicated in SEQ ID NO. 30;
(p) gene products (probable phosphate butyryltransferase;
E.C. 2.3.1.19) involved in the synthesis of butanol and/or butyric acid and having an amino acid sequence which shows at least 69% identity to the amino acid sequence indicated in SEQ ID NO. 32;
(q) gene products (probable butyrate kinase;
E.C. 2.7.2.7) involved in the synthesis of butanol and/or butyric acid and having an amino acid sequence which shows at least 84% identity to the amino acid sequence indicated in SEQ ID NO 34;
(r) gene products AcsA (acetyl-coenzyme A synthetase;
E.C. 6.2.1.1) involved in the synthesis of propionic acid and having an amino acid sequence which shows at least 85% identity to the amino acid sequence indicated in SEQ ID NO. 36;
(s) gene products Ytcl (acetate-CoA ligase;
E.C. 6.2.1.1) involved in the synthesis of propionic acid and having an amino acid sequence which shows at least 77% identity to the amino acid sequence indicated in SEQ ID NO. 38;
(t) gene products SpeA (lysine and/or arginine decarboxylase;
E.C. 4.1.1.18 or E.C. 4.1.1.19) involved in the synthesis of cadaverine and/or putrescine and having an amino acid sequence which shows at least 66% identity to the amino acid sequence indicated in SEQ ID NO. 40;
(u) gene products YsiB (probable enoyl-CoA hydratase;
E.C. 4.2.1.17) involved in the synthesis of isovaleric acid, 2-methylbutyric acid and/or isobutyric acid and having an amino acid sequence which shows at least 77% identity to the amino acid sequence indicated in SEQ ID NO. 42;
(v) gene products (similar to 3-hydroxyacyl-CoA dehydrogenase) involved in the synthesis of 2-methylbutyric acid and having an amino acid sequence which shows at least 80% identity to the amino acid sequence indicated in SEQ ID NO. 44;
(w) gene products (2-oxoglutarate dehydrogenase E1 component;
E.C. 1.2.4.2) involved in the synthesis of isovaleric acid, 2-methylbutyric acid and/or isobutyric acid and having an amino acid sequence which shows at least 82% identity to the amino acid sequence indicated in SEQ ID NO. 46;
(x) gene products YhfL (probable acid-CoA ligase;
E.C. 6.2.1.-) involved in the synthesis of propionic acid and having an amino acid sequence which shows at least 76% identity to the amino acid sequence indicated in SEQ ID NO. 48; and
(y) gene products YwhG (agmatinase;
E.C. 3.5.1.11) involved in the synthesis of cadaverine and/or putrescine and having an amino acid sequence which shows at least 97% identity to the amino acid sequence indicated in SEQ ID NO. 50. - View Dependent Claims (4)
-
-
5. A process for fermenting a substrate using a microorganism, the process comprising:
-
(a) providing a microorganism, the microorganism having at least one gene on a metabolic pathway for synthesizing a predetermined product being functionally inactivated, the metabolic pathway being selected from the group consisting of;
(1) the metabolic pathway for synthesizing isovaleric acid (as part of leucine catabolism);
(2) the metabolic pathway for synthesizing 2-methylbutyric acid and/or isobutyric acid (as part of valine and/or isoleucine catabolism) (3) the metabolic pathway for synthesizing butanol and/or butyric acid (as part of butyric acid metabolism);
(4) the metabolic pathway for synthesizing propionic acid (as part of propionate metabolism); and
(5) the metabolic pathway for synthesizing cadaverine and/or putrescine (as parts of lysine and/or arginine catabolism);
(b) providing a fermentable substrate; and
(c) permitting the microorganism to ferment the substrate. - View Dependent Claims (6, 7, 8, 9, 10, 11, 12, 13, 14, 15)
-
-
16. A process for fermenting a substrate using a microorganism, the process comprising:
-
(a) providing a microorganism, the microorganism having at least one gene on a metabolic pathway for synthesizing a predetermined product being functionally inactivated, the functionally inactivated enzyme being selected from the group consisting of;
(1) functionally inactivated enzymes of the metabolic pathway for synthesizing isovaleric acid (as part of leucine catabolism) selected from the group consisting of;
(i) L-leucine dehydrogenase (E.C. 1.4.1.9), (ii) 3-methyl-2-oxobutanoate dehydrogenase or 2-oxoglutarate dehydrogenase E1 (E.C. 1.2.4.2), (iii) enzyme for hydrolyzing isovaleryl-CoA to isovaleric acid and coenzyme A, (iv) acyl-CoA dehydrogenase (E.C. 1.3.99.-), (v) methylcrotonyl carboxylase, (vi) 3-methylglutaconyl-CoA hydratase, and (vii) enoyl-CoA hydratase (protein) (E.C. 4.2.1.17);
(2) functionally inactivated enzymes of the metabolic pathway for synthesizing 2-methylbutyric acid and/or isobutyric acid (as part of valine and/or isoleucine catabolism) from the group consisting of;
(i) branched-chain amino acid aminotransferase (E.C. 2.6.1.42), (ii) 3-methyl-2-oxobutanoate dehydrogenase or 2-oxoglutarate dehydrogenase E1 (E.C. 1.2.4.2), (iii) enzyme for hydrolyzing 2-methylbutyryl-CoA to 2-methylbutyric acid or isobutyryl-CoA to isobutyric acid and coenzyme A, (iv) acyl-CoA dehydrogenase (E.C. 1.3.99.-), (v) enoyl-CoA hydratase (protein) (E.C. 4.2.1.17), (vi) 3-hydroxy-acyl-CoA dehydrogenase (E.C. 1.1.1.35), (vii) acetyl-CoA acyltransferase, (viii) enoyl-(3-hydroxyisobutyryl)-CoA hydrolase protein and (ix) 3-hydroxyisobutyrate dehydrogenase (E.C. 1.1.1.31) or oxidoreductase (E.C.1.1.-.-);
(3) fuctionally inactivated enzymes of the metabolic pathway for synthesizing butanol and/or butyric acid (as part of butyric acid metabolism) from the group consisting of;
(i) 3-hydroxybutyryl-CoA dehydrogenase (E.C. 1.1.1.157), (ii) 3-hydroxybutyryl-CoA dehydratase (E.C. 4.2.1.55), (iii) butyryl-CoA dehydrogenase (E.C. 1.3.99.25), (iv) phosphate butyryltransferase (E.C. 2.3.1.19), (v) butyrate kinase (E.C. 2.7.2.7), (vi) butyraldehyde dehydrogenase and (vii) NADH-dependent butanol dehydrogenase A (E.C. 1.1.1.-) (4) functionally inactivated enzymes of the metabolic pathway for synthesizing propionic acid (as part of propionate metabolism) from the group consisting of;
(i) 3-hydroxybutyryl-CoA dehydrogenase (E.C. 1.1.1.157), (ii) 3-hydroxybutyryl-CoA dehydratase (E.C. 4.2.1.55), (iii) butyryl-CoA dehydrogenase (E.C. 1.3.99.25), (iv) phosphate butyryltransferase (E.C. 2.3.1.19), (v) butyrate kinase (E.C. 2.7.2.7), (vi) butyraldehyde dehydrogenase, and (viii) NADH-dependent butanol dehydrogenase A (E.C. 1.1.1.-) and (5) functionally inactivated enzymes of metabolic pathway for synthesizing cadaverine and/or putrescine (as parts of lysine and/or arginine catabolism) from the group consisting of;
(i) lysine decarboxylase (E.C. 4.1.1.18) and/or arginine decarboxylase (E.C. 4.1.1.19), (ii) agmatinase (E.C. 3.5.1.11) and (iii) ornithine decarboxylase (E.C. 4.1.1.17);
(b) providing a fermentable substrate; and
(c) permitting the microorganism to ferment the substrate.
-
-
17. A process for fermenting a substrate using a microorganism, the process comprising:
-
(a) providing a microorganism, the microorganism producing at least one functionally inactivated enzyme, the at least one functionally inactivated enzyme being a homolog, naturally active in the microorganism, to one of the proteins from B. licheniformis DSM 13 selected from the group consisting of;
(1) proteins of the metabolic pathway for synthesizing isovaleric acid (as part of leucine catabolism) selected from the group consisting of;
(i) 3-methyl-2-oxobutanoate dehydrogenase or 2-oxoglutarate dehydrogenase E1 (E.C. 1.2.4.2) defined by SEQ ID NO. 46, (ii) a subunit of acyl-CoA dehydrogenase (E.C. 1.3.99.-) defined by SEQ ID NO. 10;
(iii) a subunit of acyl-CoA dehydrogenase (E.C. 1.3.99.-) defined by SEQ ID NO. 12;
(iv) a subunit of acyl-CoA dehydrogenase (E.C. 1.3.99.-) defined by SEQ ID NO. 14;
(v) enoyl-CoA hydratase (protein) (E.C. 4.2.1.17) as defined by SEQ ID NO. 16;
(vi) enoyl-CoA hydratase (protein) (E.C. 4.2.1.17) as defined by SEQ ID NO. 18;
(vii) enoyl-CoA hydratase (protein) (E.C. 4.2.1.17) as defined by SEQ ID NO. 20; and
(viii) enoyl-CoA hydratase (protein) (E.C. 4.2.1.17) as defined by SEQ ID NO. 42;
(2) proteins of the metabolic pathway for synthesizing 2-methylbutyric acid and/or isobutyric acid (as part of valine and/or isoleucine catabolism) from the group consisting of;
(i) branched-chain amino acid aminotransferase (E.C. 2.6.1.42) defined by SEQ ID NO. 2 or 4, (ii) 3-methyl-2-oxobutanoate dehydrogenase or 2-oxoglutarate dehydrogenase E1 (E.C. 1.2.4.2) defined by SEQ ID NO. 46, (iii) acyl-CoA dehydrogenase (E.C. 1.3.99.-) defined by SEQ ID NO. 10, (iv) acyl-CoA dehydrogenase (E.C. 1.3.99.-) defined by SEQ ID NO. 12, (v) acyl-CoA dehydrogenase (E.C. 1.3.99.-) defined by SEQ ID NO. 22, (vi) acyl-CoA dehydrogenase (E.C. 1.3.99.-) defined by SEQ ID NO. 28, (vii) enoyl-CoA hydratase (protein) (E.C. 4.2.1.17) defined by SEQ ID NO. 16, (viii) enoyl-CoA hydratase (protein) (E.C. 4.2.1.17) defined by SEQ ID NO. 18, (ix) enoyl-CoA hydratase (protein) (E.C. 4.2.1.17) defined by SEQ ID NO. 20, (x) enoyl-CoA hydratase (protein) (E.C. 4.2.1.17) defined by SEQ ID NO. 42, (xi) 3-hydroxy-acyl-CoA dehydrogenase (E.C. 1.1.1.35) defined by SEQ ID NO. 44, (xii) enoyl-(3-hydroxyisobutyryl)-CoA hydrolase protein defined by SEQ ID NO. 18; and
(xiii) 3-hydroxyisobutyrate dehydrogenase (E.C. 1.1.1.31) or oxidoreductase (E.C.1.1.-.-) defined by SEQ ID NO. 30; and
(3) proteins of the metabolic pathway for synthesizing butanol and/or butyric acid (as part of butyric acid metabolism) from the group consisting of;
(i) 3-hydroxybutyryl-CoA dehydrogenase (E.C. 1.1.1.157) defined by SEQ ID NO. 14;
(ii) 3-hydroxybutyryl-CoA dehydratase (E.C. 4.2.1.55) defined by SEQ ID NO. 26;
(iii) butyryl-CoA dehydrogenase (E.C. 1.3.99.25) defined by SEQ ID NO. 10;
(iv) butyryl-CoA dehydrogenase (E.C. 1.3.99.25) defined by SEQ ID NO. 12;
(v) butyryl-CoA dehydrogenase (E.C. 1.3.99.25) defined by SEQ ID NO. 28;
(vi) phosphate butyryltransferase (E.C. 2.3.1.19) defined by SEQ ID NO. 32;
(vii) butyrate kinase (E.C. 2.7.2.7) defined by SEQ ID NO. 34; and
(viii) NADH-dependent butanol dehydrogenase A (E.C. 1.1.1.-) defined by SEQ ID NO. 8;
(4) proteins of the metabolic pathway for synthesizing propionic acid (as part of propionate metabolism) from the group consisting of;
(i) acetate-CoA ligase or synthetase or propionate-CoA ligase or synthetase (E.C. 6.2.1.1) defined by SEQ ID NO. 24;
(ii) acetate-CoA ligase or synthetase or propionate-CoA ligase or synthetase (E.C. 6.2.1.1) defined by SEQ ID NO. 36;
(iii) acetate-CoA ligase or synthetase or propionate-CoA ligase or synthetase (E.C. 6.2.1.1) defined by SEQ ID NO. 38; and
(iv) acetate-CoA ligase or synthetase or propionate-CoA ligase or synthetase (E.C. 6.2.1.1) defined by SEQ ID NO. 48; and
(5) proteins of the metabolic pathway for synthesizing cadaverine and/or putrescine (as parts of lysine and/or arginine catabolism) from the group consisting of;
(i) lysine and/or arginine decarboxylase (E.C. 4.1.1.18 or E.C. 4.1.1.19) defined by SEQ ID NO. 6;
(ii) lysine and/or arginine decarboxylase (E.C. 4.1.1.18 or E.C. 4.1.1.19) defined by SEQ ID NO. 40; and
(iii) agmatinase (E.C. 3.5.1.11) defined by SEQ ID NO. 50;
(b) providing a fermentable substrate; and
(c) permitting the microorganism to ferment the substrate.
-
-
18. A process for fermenting a substrate using a microorganism, the process comprising:
-
(a) providing a microorganism, the microorganism having at least one gene on a metabolic pathway for synthesizing a predetermined product being functionally inactivated, the enzyme being functionally inactivated at the genetic level by inactivation of a gene which corresponds to the nucleic acid which codes for a protein from B. licheniformis DSM 13 selected from the group consisting of;
(1) proteins of the metabolic pathway for synthesizing isovaleric acid (as part of leucine catabolism) selected from the group consisting of;
(i) 3-methyl-2-oxobutanoate dehydrogenase or 2-oxoglutarate dehydrogenase E1 (E.C. 1.2.4.2) coded by the nucleic acid defined by SEQ ID NO. 45;
(ii) a subunit of acyl-CoA dehydrogenase (E.C. 1.3.99.-) coded by the nucleic acid defined by SEQ ID NO. 9;
(iii) a subunit of acyl-CoA dehydrogenase (E.C. 1.3.99.-) coded by the nucleic acid defined by SEQ ID NO. 11;
(iv) a subunit of acyl-CoA dehydrogenase (E.C. 1.3.99.-) coded by the nucleic acid defined by SEQ ID NO. 21;
(v) a subunit of acyl-CoA dehydrogenase (E.C. 1.3.99.-) coded by the nucleic acid defined by SEQ ID NO. 27 (yusJ gene);
(vi) enoyl-CoA hydratase (protein) (E.C. 4.2.1.17) coded by the nucleic acid as defined by SEQ ID NO. 15;
(vii) enoyl-CoA hydratase (protein) (E.C. 4.2.1.17) coded by the nucleic acid defined by SEQ ID NO. 17;
.(viii) enoyl-CoA hydratase (protein) (E.C. 4.2.1.17) coded by the nucleic acid defined by SEQ ID NO. 19 (echA8 gene); and
(ix) enoyl-CoA hydratase (protein) (E.C. 4.2.1.17) coded by the nucleic acid defined by SEQ ID NO. 41 (ysiB gene);
(2) proteins of metabolic pathway for synthesizing 2-methylbutyric acid and/or isobutyric acid (as part of valine and/or isoleucine catabolism) from the group consisting of;
(i) branched-chain amino acid aminotransferase (E.C. 2.6.1.42) coded by the nucleic acid defined by SEQ ID NO. 1;
(ii) branched-chain amino acid aminotransferase (E.C. 2.6.1.42) coded by the nucleic acid defined by SEQ ID NO. 3;
(iii) 3-methyl-2-oxobutanoate dehydrogenase or 2-oxoglutarate dehydrogenase E1 (E.C. 1.2.4.2) coded by the nucleic acid defined by SEQ ID NO. 45, (iv) acyl-CoA dehydrogenase (E.C. 1.3.99.-) coded by the nucleic acid defined by SEQ ID NO. 9 (yusJ gene), (v) acyl-CoA dehydrogenase (E.C. 1.3.99.-) coded by the nucleic acid defined by SEQ ID NO. 11 (yusJ gene), (vi) acyl-CoA dehydrogenase (E.C. 1.3.99.-) coded by the nucleic acid defined by SEQ ID NO. 21 (yusJ gene), (vii) acyl-CoA dehydrogenase (E.C. 1.3.99.-) coded by the nucleic acid defined by SEQ ID NO. 27 (yusJ gene), (viii) enoyl-CoA hydratase (protein) (E.C. 4.2.1.17) defined by SEQ ID NO. 15(echA8 gene);
(ix) enoyl-CoA hydratase (protein) (E.C. 4.2.1.17) defined by SEQ ID NO. 17 (echA8 gene);
(x) enoyl-CoA hydratase (protein) (E.C. 4.2.1.17) defined by SEQ ID NO. 19 (echA8 gene);
(xi) enoyl-CoA hydratase (protein) (E.C. 4.2.1.17) defined by SEQ ID NO. 41 (ysiB gene);
(xii) 3-hydroxy-acyl-CoA dehydrogenase (E.C. 1.1.1.35) coded by the nucleic acid defined by SEQ ID NO. 43, (xiii) enoyl-(3-hydroxyisobutyryl)-CoA hydrolase protein coded by the nucleic acid defined by SEQ ID NO. 17; and
(xiv) 3-hydroxyisobutyrate dehydrogenase (E.C. 1.1.1.31) or oxidoreductase (E.C.1.1.-.-) coded by the nucleic acid defined by SEQ ID NO. 29 (ykwC gene);
(3) proteins of metabolic pathway for synthesizing butanol and/or butyric acid (as part of butyric acid metabolism) from the group consisting of;
(i) 3-hydroxybutyryl-CoA dehydrogenase (E.C. 1.1.1.157) coded by the nucleic acid defined by SEQ ID NO. 13;
(ii) 3-hydroxybutyryl-CoA dehydratase (E.C. 4.2.1.55) coded by the nucleic acid defined by SEQ ID NO. 25 (yngF gene);
(iii) butyryl-CoA dehydrogenase (E.C. 1.3.99.25) coded by the nucleic acid defined by SEQ ID NO. 9 (yusJ gene);
(iv) butyryl-CoA dehydrogenase (E.C. 1.3.99.25) coded by the nucleic acid defined by SEQ ID NO. 11 (yusJ gene);
(v) butyryl-CoA dehydrogenase (E.C. 1.3.99.25) coded by the nucleic acid defined by SEQ ID NO. 27 (yusJ gene);
(vi) phosphate butyryltransferase (E.C. 2.3.1.19) coded by the nucleic acid defined by SEQ ID NO. 31;
(vii) butyrate kinase (E.C. 2.7.2.7) coded by the nucleic acid defined by SEQ ID NO. 33; and
(viii) NADH-dependent butanol dehydrogenase A (E.C. 1.1.1.-) coded by the nucleic acid defined by SEQ ID NO. 7 (yugJ gene);
(4) proteins of a metabolic pathway for synthesizing propionic acid (as part of propionate metabolism) from the group consisting of;
(i) acetate-CoA ligase or synthetase or propionate-CoA ligase or synthetase (E.C. 6.2.1.1) coded by the nucleic acid defined by SEQ ID NO. 35 (acsA gene);
(ii) acetate-CoA ligase or synthetase or propionate-CoA ligase or synthetase (E.C. 6.2.1.1) coded by the nucleic acid defined by SEQ ID NO. 37 (ytcl gene);
(iii) acetate-CoA ligase or synthetase or propionate-CoA ligase or synthetase (E.C. 6.2.1.1) coded by the nucleic acid defined by SEQ ID NO. 47 (yhfL gene);
(iv) acetate-CoA ligase or synthetase or propionate-CoA ligase or synthetase (E.C. 6.2.1.1) coded by the nucleic acid defined by SEQ ID NO. 23 (acsA gene);
(5) proteins of a metabolic pathway for synthesizing cadaverine and/or putrescine (as parts of lysine and/or arginine catabolism) from the group consisting of;
(i) lysine and/or arginine decarboxylase (E.C. 4.1.1.18 or E.C. 4.1.1.19) coded by the nucleic acid defined by SEQ ID NO. 5 (speA gene);
(ii) lysine and/or arginine decarboxylase (E.C. 4.1.1.18 or E.C. 4.1.1.19) coded by the nucleic acid defined by SEQ ID NO. 39 (speA gene); and
(iii) agmatinase (E.C. 3.5.1.11) coded by the nucleic acid defined by SEQ ID NO. 49 (ywhG gene);
(b) providing a fermentable substrate; and
(c) permitting the microorganism to ferment the substrate.
-
-
19. A process for functionally inactivating at least one gene of a microorganism, the process comprising
(a) providing at least one predetermined nucleic acid selected from the group consisting of: -
(1) nucleic acids coding for a gene product (putative branched-chain amino acid aminotransferase;
E.C. 2.6.1.42) involved in the synthesis of 2-methylbutyric acid and/or isobutyric acid and having a nucleotide sequence which shows at least 67% identity to the nucleotide sequence indicated in SEQ ID NO. 1;
(2) nucleic acids coding for a gene product (putative branched-chain amino acid aminotransferase;
E.C. 2.6.1.42) involved in the synthesis of 2-methylbutyric acid and/or isobutyric acid and having a nucleotide sequence which shows at least 78% identity to the nucleotide sequence indicated in SEQ ID NO. 3;
(3) nucleic acids speA coding for a gene product (lysine and/or arginine decarboxylase;
E.C. 4.1.1.18 or 4.1.1.19) involved in the synthesis of cadaverine and/or putrescine and having a nucleotide sequence which shows at least 78% identity to the nucleotide sequence indicated in SEQ ID No. 5;
(4) nucleic acids yugJ coding for a gene product (NADH-dependent butanol dehydrogenase A;
E.C. 1.1.1.-) involved in the synthesis of butanol and/or butyric acid and having a nucleotide sequence which shows at least 81% identity to the nucleotide sequence indicated in SEQ ID NO. 7;
(5) nucleic acids coding for a gene product (acyl-CoA dehydrogenase;
E.C. 1.3.99.-) involved in the synthesis of isovaleric acid, 2-methylbutyric acid, isobutyric acid, butanol and/or butyric acid and having a nucleotide sequence which shows at least 79% identity to the nucleotide sequence indicated in SEQ ID NO. 9;
(6) nucleic acids coding for a gene product (acyl-CoA dehydrogenase;
E.C. 1.3.99.-) involved in the synthesis of isovaleric acid, 2-methylbutyric acid, isobutyric acid, butanol and/or butyric acid and having a nucleotide sequence which shows at least 64% identity to the nucleotide sequence indicated in SEQ ID NO. 11;
(7) nucleic acids coding for a gene product (3-hydroxybutyryl-CoA dehydrogenase;
E.C. 1.1.1.157) involved in the synthesis of butanol and/or butyric acid and having a nucleotide sequence which shows at least 67% identity to the nucleotide sequence indicated in SEQ ID NO. 13;
(8) nucleic acids coding for a gene product (putative enoyl-Coa hydratase protein;
E.C. 4.2.1.17) involved in the synthesis of isovaleric acid, 2-methylbutyric acid and/or isobutyric acid and having a nucleotide sequence which shows at least 65% identity to the nucleotide sequence indicated in SEQ ID NO. 15;
(9) nucleic acids coding for a gene product (probable enoyl-(3-hydroxyisobutyryl)-coenzyme A hydrolase protein) involved in the synthesis of isovaleric acid, 2-methylbutyric acid and/or isobutyric acid and having a nucleotide sequence which shows at least 66% identity to the nucleotide sequence indicated in SEQ ID NO. 17;
(10) nucleic acids echA8 coding for a gene product (probable enoyl-CoA hydratase;
E.C. 4.2.1.17) involved in the synthesis of isovaleric acid, 2-methylbutyric acid and/or isobutyric acid and having a nucleotide sequence which shows at least 48% identity to the nucleotide sequence indicated in SEQ ID NO. 19;
(11) nucleic acids coding for a gene product (acyl-CoA dehydrogenase;
E.C. 1.3.99.-) involved in the synthesis of isovaleric acid, 2-methylbutyric acid and/or isobutyric acid and having a nucleotide sequence which shows at least 54% identity to the nucleotide sequence indicated in SEQ ID NO. 21;
(12) nucleic acids acsA coding for a gene product (acetyl-coenzyme A synthetase;
E.C. 6.2.1.1) involved in the synthesis of propionic acid and having a nucleotide sequence which shows at least 67% identity to the nucleotide sequence indicated in SEQ ID NO. 23;
(13) nucleic acids yngF coding for a gene product (3-hydroxybutyryl-CoA dehydratase;
E.C. 4.2.1.55) involved in the synthesis of butanol and/or butyric acid and having a nucleotide sequence which shows at least 68% identity to the nucleotide sequence indicated in SEQ ID NO. 25;
(14) nucleic acids yusJ coding for a gene product (acyl-CoA dehydrogenase;
E.C. 1.3.99.-) involved in the synthesis of isovaleric acid, 2-methylbutyric acid, isobutyric acid, butanol and/or butyric acid and having a nucleotide sequence which shows at least 77% identity to the nucleotide sequence indicated in SEQ ID NO. 27;
(15) nucleic acids ykwC coding for a gene product (hypothetical oxidoreductase;
E.C. 1.1.-.-) involved in the synthesis of 2-methylbutyric acid and/or isobutyric acid and having a nucleotide sequence which shows at least 77% identity to the nucleotide sequence indicated in SEQ ID NO. 29;
(16) nucleic acids coding for a gene product (probable phosphate butyryltransferase;
E.C. 2.3.1.19) involved in the synthesis of butanol and/or butyric acid and having a nucleotide sequence which shows at least 51% identity to the nucleotide sequence indicated in SEQ ID NO. 31;
(17) nucleic acids coding for a gene product (probable butyrate kinase;
E.C. 2.7.2.7) involved in the synthesis of butanol and/or butyric acid and having a nucleotide sequence which shows at least 77% identity to the nucleotide sequence indicated in SEQ ID NO. 33;
(18) nucleic acids acsA coding for a gene product (acetyl-coenzyme A synthetase;
E.C. 6.2.1.1) involved in the synthesis of propionic acid and having a nucleotide sequence which shows at least 79% identity to the nucleotide sequence indicated in SEQ ID NO. 35;
(19) nucleic acids ytcl coding for a gene product (acetate-CoA ligase;
E.C. 6.2.1.1) involved in the synthesis of propionic acid and having a nucleotide sequence which shows at least 74% identity to the nucleotide sequence indicated in SEQ ID NO. 37;
(20) nucleic acids speA coding for a gene product (lysine and/or arginine decarboxylase;
E.C. 4.1.1.18 or E.C. 4.1.1.19) involved in the synthesis of cadaverine and/or putrescine and having a nucleotide sequence which shows at least 68% identity to the nucleotide sequence indicated in SEQ ID NO. 39;
(21) nucleic acids ysiB coding for a gene product (probable Enoyl-CoA hydratase;
E.C. 4.2.1.17) involved in the synthesis of isovaleric acid, 2-methylbutyric acid and/or isobutyric acid and having a nucleotide sequence which shows at least 75% identity to the nucleotide sequence indicated in SEQ ID NO. 41;
(22) nucleic acids coding for a gene product (similar to 3-hydroxyacyl-CoA dehydrogenase;
E.C. 1.1.1.35) involved in the synthesis of 2-methylbutyric acid and having a nucleotide sequence which shows at least 76% identity to the nucleotide sequence indicated in SEQ ID NO. 43;
(23) nucleic acids coding for a gene product (2-oxoglutarate dehydrogenase E1 component;
E.C. 1.2.4.2) involved in the synthesis of isovaleric acid, 2-methylbutyric acid and/or isobutyric acid and having a nucleotide sequence which shows at least 80% identity to the nucleotide sequence indicated in SEQ ID NO. 45;
(24) nucleic acids yhfL coding for a gene product (probable acid-CoA ligase;
E.C. 6.2.1.-) involved in the synthesis of propionic acid and having a nucleotide sequence which shows at least 67% identity to the nucleotide sequence indicated in SEQ ID NO. 47; and
(25) nucleic acids ywhG coding for a gene product (agmatinase;
E.C. 3.5.1.11) involved in the synthesis of cadaverine and/or putrescine and having a nucleotide sequence which shows at least 85% identity to the nucleotide sequence indicated in SEQ ID NO. 49.(b) constructing a vector with the at least one predetermined nucleic acid; and
(c) introducing the vector into the target microorganism. - View Dependent Claims (20, 21)
-
-
22. A microorganism in which at least one of the genes which corresponds to the nucleic acid which codes for one predetermined protein of B. licheniformis DSM 13 is functionally inactivated, the predetermined protein being selected from the group consisting of:
-
putative branched-chain amino acid aminotransferase (E.C. 2.6.1.42) defined by SEQ ID NO. 1, putative branched-chain amino acid aminotransferase (E.C. 2.6.1.42) defined by SEQ ID NO. 3, lysine and/or arginine decarboxylase (protein SpeA;
E.C. 4.1.1.18 or E.C. 4.1.1.19) defined by SEQ ID NO. 5 (speA gene),NADH-dependent butanol dehydrogenase A (protein YugJ;
E.C. 1.1.1.-) defined by SEQ ID NO. 7 (yugJ gene),butyryl-CoA dehydrogenase (E.C. 1.3.99.25) or acyl-CoA dehydrogenase (E.C. 1.3.99.-) defined by SEQ ID NO. 9, butyryl-CoA dehydrogenase (E.C. 1.3.99.25) or acyl-CoA dehydrogenase (E.C. 1.3.99.-) defined by SEQ ID NO. 11, 3-hydroxybutyryl-CoA dehydrogenase (E.C. 1.1.1.157) defined by SEQ ID NO. 13, putative enoyl-CoA hydratase protein (E.C. 4.2.1.17) defined by SEQ ID NO. 15, probable enoyl-(3-hydroxyisobutyryl)-CoA hydrolase protein (E.C.4.2.1.17) defined by SEQ ID NO. 17, probable enoyl-CoA hydratase (protein EchA8;
E.C. 4.2.1.17) defined by SEQ ID NO. 19 (echA8 gene),acyl-CoA dehydrogenase (E.C. 1.3.99.-) defined by SEQ ID NO. 21, acetate-CoA ligase or propionate-CoA ligase (or synthetase;
protein AcsA;
E.C. 6.2.1.1) defined by SEQ ID NO. 23 (acsA gene),3-hydroxybutyryl-CoA dehydratase (protein YngF;
E.C. 4.2.1.55) defined by SEQ ID NO. 25 (yngF gene),butyryl-CoA dehydrogenase (protein YusJ;
E.C. 1.3.99.25) or acyl-CoA dehydrogenase (E.C. 1.3.99.-) defined by SEQ ID NO. 27 (yusJ gene),3-hydroxyisobutyrate dehydrogenase (protein YkwC;
E.C. 1.1.1.31) or oxidoreductase (E.C.1.1.-.-) defined by SEQ ID NO. 29 (ykwC gene),probable phosphate butyryltransferase (E.C. 2.3.1.19) defined by SEQ ID NO. 31, probable butyrate kinase (E.C. 2.7.2.7) defined by SEQ ID NO. 33, acetate-CoA ligase or synthetase or propionate-CoA ligase or synthetase (protein AcsA;
E.C. 6.2.1.1) defined by SEQ ID NO. 35 (acsA gene),acetate-CoA ligase or propionate-CoA ligase (protein Ytcl;
E.C. 6.2.1.1) defined by SEQ ID NO. 37 (ytcl gene),lysine and/or arginine decarboxylase (protein speA;
E.C. 4.1.1.18 or E.C. 4.1.1.19) defined by SEQ ID NO. 39 (speA gene),probable enoyl-CoA hydratase (E.C. 4.2.1.17) defined by SEQ ID NO. 41 (ysiB gene), similar to 3-hydroxyacyl-CoA dehydrogenase (E.C. 1.1.1.35) defined by SEQ ID NO. 43, 3-methyl-2-oxobutanoate dehydrogenase or 2-oxoglutarate dehydrogenase E1 (E.C. 1.2.4.2) defined by SEQ ID NO. 45, probable acetate-CoA ligase or propionate-CoA ligase (protein YhfL;
E.C. 6.2.1.1) or acid-CoA ligase (E.C. 6.2.1.-) defined by SEQ ID NO. 47 (yhfL gene) oragmatinase (E.C. 3.5.1.11) defined by SEQ ID NO. 49 (ywhG gene). - View Dependent Claims (23, 24, 25, 26, 27, 28, 29, 30)
-
Specification