Methods for the production of methacrylates

1. A method for the production of a methacrylate comprising

a) providing a genetically modified microorganism with increased tolerance to a C 3 -C 12 methacrylate ester compared to the wild type microorganism in a fermentation medium and

b) growing the microorganism under conditions whereby a C 3 -C 12 methacrylate ester is produced;

wherein the microorganism is E coli;

wherein the genetically modified microorganism comprises a mutation in a wild type soxR nucleic acid sequence;

wherein the wild type soxR nucleic acid sequence comprises SEQ ID NO. 1 or a homologue thereof, and

wherein the mutant soxR nucleic acid sequence encodes a mutant SoxR protein comprising R20 substituted with H.

2. The method according to claim 1 wherein the microorganism is tolerant to at least 20% v/v C 3 -C 12 methacrylate ester when grown in a liquid medium at about 37° C.

3. The method according to claim 1 wherein the genetically modified microorganism further comprises a mutation in one or more of:

an acrR nucleic acid sequence comprising SEQ ID NO. 3 or a homologue thereof, a rob nucleic acid sequence comprising SEQ ID NO. 5 or a homologue thereof, a marR nucleic acid sequence comprising SEQ ID NO. 7 or a homologue thereof.

4. The method according to claim 3 , wherein the mutant acrR nucleic acid sequence encodes a mutant AcrR protein, and wherein the mutation is selected from one of the following: a substitution of V29 with another amino acid, a frameshift mutation at T32, A191 or position 49 in AcrR.

5. A method for growing or maintaining a microorganism in the presence of a C 3 -C 12 methacrylate ester comprising providing a genetically modified microorganism with increased tolerance to a C 3 -C 12 methacrylate ester compared to a wild type microorganism in a fermentation medium under conditions whereby a C 3 -C 12 methacrylate ester is produced,

wherein the microorganism is E coli;

wherein the genetically modified microorganism comprises a mutation is in a wild type soxR nucleic acid sequence;

wherein the wild type soxR nucleic acid sequence comprises SEQ ID NO. 1 or a homologue thereof; and

wherein the mutant soxR nucleic acid sequence encodes a mutant SoxR protein comprising R20 substituted with H.

6. A fermentation medium comprising a C 3 -C 12 methacrylate ester and a genetically modified microorganism with increased tolerance to a C 3 -C 12 methacrylate ester compared to a wild type microorganism;

wherein the microorganism is E. coli;

wherein the genetically modified microorganism comprises a mutation is in a wild type soxR nucleic acid sequence;

wherein the wild type soxR nucleic acid sequence comprises SEQ ID NO. 1 or a homologue thereof; and

wherein the mutant soxR nucleic acid sequence encodes a mutant SoxR protein comprising R20 substituted with H.

7. A method for the isolation of a methacrylate tolerant microorganism comprising:

a) providing a microorganism in a fermentation medium

b) contacting the microorganism with a methacrylate; and

c) isolating the viable microorganism of step (b)

wherein the viable microorganism is tolerant to at least 20% v/v a methacrylate when grown in liquid medium at about 37° C.

wherein the microorganism is E coli;

wherein the microorganism comprises a mutation in a wild type soxR nucleic acid sequence;

wherein the wild type soxR nucleic acid sequence comprises SEQ ID NO. 1 or a homologue thereof; and

wherein the mutant soxR nucleic acid sequence encodes a mutant SoxR protein comprising R20 substituted with H.

8. The method according to claim 3 , wherein the mutant rob nucleic acid sequence encodes a mutant Rob protein, and wherein the mutation is selected from one of the following: a substitution of A70 or a substitution of R156 with another amino acid in Rob.

9. The method according to claim 3 , wherein the mutant marR nucleic acid sequence encodes a mutant MarR protein, and wherein the mutation is a substitution of V84 with another amino acid in MarR.