Recombinant production systems for aromatic molecules

1. A modified recombinant host cell for producing a phenolic compound, comprising

i) a first expression system that comprises at least one nucleotide sequence that encodes a 6-methylsalicylic acid synthase (6-MSAS) or an orsellinic acid synthase (OSAS), capable of being expressed, and

ii) a second expression system that comprises at least one nucleotide sequence that encodes a holo acyl carrier protein (ACP) synthase, wherein the ACP synthase pantetheinylates said 6-MSAS or OSAS, and

iii) a third expression system that comprises at least one nucleotide sequence that encodes a functional decarboxylase which catalyzes decarboxylation to form said phenolic compound, such that the product isolated from said cell is a phenolic compound, wherein either:

the first expression system for 6-MSAS or OSAS and the second expression system for ACP synthase are present on a single vector or on separate vectors, and wherein the third expression system for a functional decarboxylase is present on the same vector as the 6-MSAS or OSAS, or the holo ACP synthase, or both, or is present on a separate vector, or;

wherein the first, second or third expression systems are integrated into the host cell chromosome or expressed from yeast artificial chromosomes (YACs).

2. The modified cell of claim 1 , wherein the phenolic compound is meta-cresol or orcinol.

3. The modified cell of claim 1 wherein said 6-MSAS is from a fungus or a prokaryote.

4. The modified cell of any one of claims 1 , 2 , and 3 , wherein the nucleotide sequence encoding the decarboxylase is selected from the group consisting of the 6-MSA decarboxylase gene from P. patulum , the PatG gene from Aspergillus clavatus , the OSA decarboxylase gene from Gliocladium roseum , the 2,3-dihydroxybenzoic acid decarboxylase genes from Aspergillus species, and the 5-carboxyvanillate decarboxylase gene from Sphingomonas paucimobilis SYK-6.

5. The modified cell of claim 1 wherein said 6-MSAS or OSAS and said holo ACP synthase are expressed from a dicistronic messenger RNA.

6. The modified cell of claim 1 wherein the host organism is selected from the group consisting of Saccharomyces cerevisiae, Pichia pastoris , and an Aspergillus species.

7. A method of producing a phenolic compound, the method comprising

a) providing a modified recombinant host cell comprising

i) a first expression system that comprises at least one nucleotide sequence encoding a 6-methylsalicylic acid synthase (6-MSAS) or an orsellinic acid synthase (OSAS), capable of being expressed;

ii) a second expression system that comprises at least one nucleotide sequence encoding a holo acyl carrier protein (ACP) synthase, wherein the ACP synthase pantetheinylates said 6-MSAS or OSAS; and

iii) a third expression system that comprises at least one nucleotide sequence that encodes a functional decarboxylase;

b) producing a first phenolic compound in the recombinant host cell; and

c) synthesizing a second phenolic compound from the first phenolic compound, wherein recombinant decarboxylase catalyzes decarboxylation of the first phenolic compound to form the second phenolic compound, wherein either:

the first expression system for 6-MSAS or OSAS and the second expression system for ACP synthase are present on a single vector or on separate vectors, and wherein the third expression system for a functional decarboxylase is present on the same vector as the 6-MSAS or OSAS, or the holo ACP synthase, or both, or is present on a separate vector, or;

wherein the first, second or third expression systems are integrated into the host cell chromosome or expressed from yeast artificial chromosomes (YACs).

8. The method of claim 7 , wherein the first phenolic compound is 6-MSA or orsellinic acid (OSA) and wherein the second phenolic compound is meta-cresol or orcinol.

9. The modified cell of claim 1 , wherein the phenolic compound is an alkylated phenolic compound and wherein the modified host cell further comprises

(iv) a fourth expression system that comprises at least one nucleotide sequence that encodes an O-methyltransferase (OMT), such that the product isolated from said cell is an alkylated phenolic compound.

10. The modified cell of claim 9 , wherein the alkylated phenolic compound is 3-methylanisole or 3,5-dimethoxytoluene.

11. The modified cell of claim 9 , wherein the OMT is encoded by a R. chinensis gene selected from the group consisting of OOMT1, OOMT2, OOMT3, OOMT4, COMT1.

12. The method of claim 7 , which is for producing an alkylated phenolic compound, the method comprising d) isolating the second phenolic compound produced in the recombinant host cell; and e) alkylating the second phenolic compound of step d) to form the alkylated phenolic compound by treating with an alkylating agent.

13. The method of claim 12 , wherein the second phenolic compound is meta-cresol or orcinol.

14. The method of any one of claims 7 , 8 , 12 , and 13 , wherein the nucleotide sequence encoding the decarboxylase is selected from the group consisting of the 6-MSA decarboxylase gene from P. patulum , the PatG gene from Aspergillus clavatus , the OSA decarboxylase gene from Gliocladium roseum , the 2,3-dihydroxybenzoic acid decarboxylase genes from Aspergillus species, and the 5-carboxyvanillate decarboxylase from Sphingomonas paucimobilis SYK-6.