Selective advantage in fermentation
Disclosed are transformed cells and related nucleotide and protein sequences, and fermentation compositions and methods, all of which are related to providing selective advantage in fermentation. For example, a selective advantage results from transformation of a cell with a nucleic acid that allows a transformed cell to metabolize one or more nitrogen-, phosphorous-, and/or sulfur-containing compounds that a native cell of the same species as the transformed cell cannot metabolize, and from fermentation of the transformed cell using one or more feedstocks, such as fractioned grain, which are depleted in or free of conventional nitrogen-, phosphorous-, and/or sulfur-containing compounds that a native cell of the same species as the transformed cell can metabolize. Also disclosed are methods for improved oxygen transfer in an aerobic or microaerobic fermentation.
1. A composition comprising
a) a fermentation mixture comprising
i) a first fraction consisting essentially of a fraction of a fractionated grain, and
ii) a second fraction comprising one or more nitrogen-containing compounds; and
b) a transformed cell comprising an exogenous nucleic acid molecule that encodes an enzyme for metabolizing the one or more nitrogen-containing compounds as a source of nitrogen; wherein the one or more nitrogen-containing compounds cannot be metabolized by a native, untransformed cell of the same species, and wherein the enzyme is a melamine deaminase (EC 3.5.4.45), a guanine deaminase (EC 3.5.4.3), a N-isopropylammelide isopropylamino (Ammelide) hydrolyase (EC 3.5.99.4), a cyanuric acid hydrolyase (EC 3.5.2.15), a biuret amidohydrolase (EC 3.5.1.84), an allophanate hydrolyase (EC 3.5.1.54), a cyanamide hydratase (EC 4.2.1.69), a urease (EC 3.5.1.5), or a urea carboxylase (EC 6.3.4.6).
2. The composition of claim 1 , wherein the one or more nitrogen-containing compounds are not naturally found in the grain.
3. The composition of claim 1 , wherein the fraction of the fractionated grain is an endosperm fraction.
4. The composition of claim 1 , wherein the grain is corn, wheat, sorghum, rye, triticale, oats, rice, millets, barley, teff, wild rice, spelt, buckwheat, amaranth, quinoa, kaniwa, or fonio.
5. The composition of claim 1 , wherein:
the second fraction comprises, in an amount from about 10% by weight to about 100% by weight, one or more nitrogen-containing compounds of any one of formulas I-III, or a salt thereof; and
the compound of formula I is
wherein, independently for each occurrence,
is a five-, six, nine-, or ten-membered aryl or heteroaryl group; R is —OH, —CO 2 H, —NO 2 , —CN, substituted or unsubstituted amino, or substituted or unsubstituted alkyl; and
n is 0, 1, 2, 3, 4, or 5;
the compound of formula II is
wherein, independently for each occurrence,
X is —NH—, —N(alkyl)—, —O—, —C(R 1 ) 2 —, —S—, or absent;
Y is —H, —NH 2 , —N(H)(alkyl), —N(alkyl) 2 , —CO 2 H, —CN, or substituted or unsubstituted alkyl; and
R 1 is —H, —OH, —CO 2 H, —NO 2 , —CN, substituted or unsubstituted amino, or substituted or unsubstituted alkyl; and
the compound of formula III is
wherein, independently for each occurrence,
Y is —H, —NH 2 , —N(H)(alkyl), —N(alkyl) 2 , —CO 2 H, —CN, or substituted or unsubstituted alkyl.
6. The composition of claim 5 , wherein the one or more nitrogen-containing compounds are selected from the group consisting of Hydrazine, 5-Aminotetrazole, Tetrazole, Melamine, Cyanamide, 2-Cyanoguanidine, Sodium azide, Carbohydrazide, 1,2,3-Triazole, 1,2,4-Triazole, 1,3-Diaminoguanidine HCl, Ammeline, 1,3,5-triazine, Aminoacetonitrile, Cyanoethylhydrazine, Azodicarbonamide, Biurea, Formamidoxime, 1,2-Dimethylhydrazine, 1,1-Dimethylhydrazine, ethylhydrazine, Ethylenediamine, Sodium dicyanamide, Guanidine carbonate, Methylamine, Ammelide, Hydroxylamine, Malononitrile, Biuret, Diethylenetriamine, Hexamethylenetetramine, Triethylenetetramine, 1,3-Diaminopropane, Triethylenetetramine, 1,3-Diaminopropane, Hydroxyurea, Tetraethylenepentamine, Thiourea, Succinonitrile, Calcium cyanamide, Cyanuric acid, Aminoethylpiperazine, Piperazine, Dimethylamine, Ethylamine, dalfampridine, Tetranitromethane, Imidazolidinyl urea, Trinitromethane, malonamide, Chloramine, Allophanate, Trimethylamine, Nitromethane, Acetaldoxime, Diazolidinyl urea, 1,2-Cyclohexanedione dioxime, Acetone oxime, Thioacetamide, Sodium thiocyanate, Isothiazole, Thiazole, Dimethylacetamide, Isothiazolinone, Methylene blue, Diethanolamine, Aspartame, Benzisothiazolinone, urea, and Acesulfame potassium.
7. The composition of claim 1 , wherein the enzyme is an allophanate hydrolase.
8. The composition of claim 1 , wherein the transformed cell is selected from the group consisting of algae, bacteria, molds, fungi, plants, and yeasts.
9. The composition of claim 8 , wherein:
the transformed cell is a bacterium;
the transformed cell comprises a second genetic modification; and
said second genetic modification confers resistance to a bacteriophage.
10. The composition of claim 9 , wherein the second genetic modification confers resistance to a bacteriophage selected from the group consisting of Wphi, Mu, T1, T2, T3, T4, T5, T6, T7, P1, P2, P4, P22, fd, phi6, phi29, phi31, phiC31, phi35, phi36, phi48, phi50, phi80, phiX174, SP01, M13, MS2, PM2, SSV-1, L5, PRD1, Qbeta, lambda, UC-1, HK97, and HK022.
11. The composition of claim 1 , wherein the transformed cell comprises one or more exogenous nucleic acid molecules that encode a melamine deaminase (EC 3.5.4.45), a guanine deaminase (EC 3.5.4.3), a N-isopropylammelide isopropylamino (Ammedlide) hydrolyase (EC 3.5.99.4), a cyanuric acid hydrolyase (EC 3.5.2.15), a biuret amidohydrolase (EC 3.5.1.84), and an allophanate hydrolyase (EC 3.5.1.54).
12. The composition of claim 1 , wherein the transformed cell comprises one or more exogenous nucleic acid molecule that encode a cyanamide hydratase (EC 4.2.1.69), a urea carboxylase (EC 6.3.4.6), and an allophanate hydrolyase (EC 3.5.1.54).
13. The composition of claim 1 , wherein the fermentation mixture does not comprise a source of nitrogen other than the one or more nitrogen-containing compounds.
14. The composition of claim 8 , wherein the yeast is an oleaginous yeast.
15. The composition of claim 8 , wherein the yeast is a Saccharomyces cerevisiae or Yarrowia lipolytica.
16. A method for fermentation, comprising incubating the composition of claim 1 , thereby producing a fermentation product and
wherein the transformed cell of the said composition is selected from the group consisting of algae, bacteria, molds, fungi, plants, and yeasts.
17. The method of claim 16 , wherein the yeast is an oleaginous yeast.
18. The method of claim 16 , wherein the yeast is a Saccharomyces cerevisiae or Yarrowia lipolytica.
19. The method of claim 16 , wherein the grain is corn, wheat, sorghum, rye, triticale, oats, rice, millets. barley, teff, wild rice, spelt, buckwheat, amaranth, quinoa, kaniwa, or fonio.
20. The method of claim 16 , wherein the one or more nitrogen-containing compounds are selected from the group consisting of Hydrazine, 5-Aminotetrazole, Tetrazole, Melamine, Cyanamide, 2-Cyanoguanidine, Sodium azide, Carbohydrazide, 1,2,3-Triazole, 1,2,4-Triazole, 1,3-Diaminoguanidine HCl, Ammeline, 1,3,5-triazine, Aminoacetonitrile, Cyanoethylhydrazine, Azodicarbonamide, Biurea, Formamidoxime, 1,2-Dimethylhydrazine, 1,1-Dimethylhydrazine, ethylhydrazine, Ethylenediamine, Sodium dicyanamide, Guanidine carbonate, Methylamine, Ammelide, Hydroxylamine, Malononitrile, Biuret, Diethylenetriamine, Hexamethylenetetramine, Triethylenetetramine, 1,3-Diaminopropane, Triethylenetetramine, 1,3-Diaminopropane, Hydroxyurea, Tetraethylenepentamine, Thiourea, Succinonitrile, Calcium cyanamide, Cyanuric acid, Aminoethylpiperazine, Piperazine, Dimethylamine, Ethylamine, dalfampridine, Tetranitromethane, Imidazolidinyl urea, Trinitromethane, malonamide, Chloramine, Allophanate, Trimethylamine, Nitromethane, Acetaldoxime, Diazolidinyl urea, 1,2-Cyclohexanedione dioxime, Acetone oxime, Thioacetamide, Sodium thiocyanate, Isothiazole, Thiazole, Dimethylacetamide, Isothiazolinone, Methylene blue, Diethanolamine, Aspartame, Benzisothiazolinone, urea, and Acesulfame potassium.