Biological alkane oxidation
The invention relates to a method for oxidizing an alkane, comprising contacting the alkane with a type alkB oxidoreductase and using a type alkB oxidoreductase to prepare a mixture of oxidation products of an alkane, wherein the ratio of carboxylic acid to alcohol in the oxidation products is preferably greater than 1:1.
1. A method for preparing a mixture of oxidation products of a C 1 -C 5 alkane comprising:
contacting a C 1 -C 5 alkane with oxygen in the presence of an AlkB oxidoreductase, wherein the amino acid sequence of said AlkB oxidoreductase is at least 90% identical to that of the AlkB oxidoreductase of Pseudomonas putida described by SEQ ID NO: 6 for a time and under conditions sufficient to oxidize the alkane to a corresponding alcohol and carboxylic acid; and
recovering a mixture of oxidation products where the ratio of carboxylic acid to alcohol is greater than 1:1.
2. The method according to claim 1 , wherein the alkane is a C 1 -C 4 alkane.
3. The method according to claim 1 , wherein the C 1 to C 5 alkane is a branched alkane.
4. The method according to claim 1 , wherein the AlkB oxidoreductase is AlkB from Pseudomonas putida GPO1.
5. The method according to claim 1 , wherein the AlkB oxidoreductase is in the form of a whole-cell catalyst.
6. The method according to claim 1 , wherein the AlkB oxidoreductase is in the form of a purified polypeptide.
7. The method according to claim 1 , comprising recovering a mixture of oxidation products where the ratio of carboxylic acid to alcohol in the oxidation products is greater than 5:1.
8. The method according to claim 3 , wherein the alkane is a C 4 or C 5 alkane.
9. The method according to claim 1 , comprising recovering a mixture of oxidation products where the ratio of carboxylic acid to alcohol is greater than 12:1.
10. The method according to claim 1 , comprising recovering a mixture of oxidation products where the ratio of carboxylic acid to alcohol is greater than 20:1.
11. The method according to claim 1 , comprising recovering a mixture of oxidation products where the ratio of carboxylic acid to alcohol is greater than 40:1.
12. The method according to claim 1 , wherein the C 1 -C 5 alkane is oxidized predominantly into a carboxylic acid.
13. The method according to claim 1 , wherein the C 1 -C 5 alkane is oxidized at a terminal carbon atom.
14. A method for preparing a mixture of oxidation products of an alkane comprising:
contacting a C 1 -C 5 alkane with oxygen and with an AlkB oxidoreductase and optionally AlkG or AlkT enzyme(s),
wherein the amino acid sequence of said AlkB oxidoreductase is at least 90% identical to that of the alkB oxidoreductase of Pseudomonas putida described by SEQ ID NO: 6, the amino acid sequence of AlkG is at least 90% identical to the AlkG sequence described by SEQ ID NO: 7, and the amino acid sequence of AlkT is at least 90% identical to the AlkT sequence described by SEQ ID NO: 8; and
recovering a mixture of oxidation products where the ratio of carboxylic acid to alcohol is greater than 1:1.
15. The method according to claim 14 , wherein the alkB oxidoreductase is expressed along with an AlkG that has a sequence that is at least 90% identical to the AlkG sequence described by SEQ ID NO: 7 and an AlkT that has a sequence that is at least 90% identical to the AlkT sequence described by SEQ ID NO: 8.
16. The method according to claim 14 , wherein the alkane is contacted with oxygen and with a Pseudomonas putida enzyme consisting of an alkB oxidoreductase that is at least 90% identical to that of the AlkB oxidoreductase of Pseudomonas putida described by SEQ ID NO: 6.
17. The method according to claim 14 , wherein said alkB oxidoreductase is contained within a whole cell catalyst or a crude lysate thereof.
18. The method according to claim 14 , wherein said alkB oxidoreductase is in a non-cellular, purified form that when resolved on an SDS gel represents at least 99% of the visible proteins.
19. The method of claim 1 , wherein the alkB oxidoreductase is a purified AlkB oxidoreductase and is in either soluble or immobilized form.
20. The method of claim 1 , wherein the AlkB oxidoreductase is a form of an engineered recombinant whole cell catalyst that expresses more AlkB oxidoreductase than a corresponding non-recombinant whole cell catalyst, wherein said engineered recombinant whole cell catalyst contains an exogenous nucleic acid sequence encoding an AlkB oxidoreductase on a plasmid or integrated into its genome.