METHOD TO PRODUCE BRANCHED-CHAIN POLYHYDROXYALKANOATES AND BRANCHED-CHAIN 3-HYDROXYACIDS FROM GLUCOSE
Disclosed herein are methods and compositions that enable the production of branched-chain polyhydroxyalkanoates, branched-chain 3-hydroxyacids (BCHA) (PHA monomers), and branched-chain fatty acids (BCFA) in Pseudomonas putida KT2440. The branched-chain molecules enabled by this platform enable novel chemistries that are not accessible via the existing paradigm which is limited to straight-chain molecules. Further disclosed herein are methods and compositions for deciphering the mechanisms of bacterial fatty acid biosynthesis is crucial for both the engineering of bacterial hosts to produce fatty acid-derived molecules and the development of new antibiotics.
1 . A non-naturally occurring Pseudomonas sp. comprising a non-naturally occurring gene encoding for malonyl-ACP decarboxylase (MadB) wherein the gene (madB) has greater than 70% sequence identity to the nucleotide sequence of PP_0262 from P. putida KT 2440.
2 . The non-naturally occurring Pseudomonas sp. of claim 1 wherein the Pseudomonas sp. is P. putida KT 2440.
3 . A method for initiating fatty acid biosynthesis in P. putida sp. comprising overexpressing a non-naturally occurring gene encoding for malonyl-ACP decarboxylase (MadB) wherein the gene (madB) has greater than 70% sequence identity to the nucleotide sequence of PP_0262 from P. putida KT 2440.
4 . A non-naturally occurring Pseudomonas sp. useful for the production of branched-chain polyhydroxyalkanoates, branched-chain 3-hydroxyacids (BCHA) (PHA monomers), and branched-chain fatty acids (BCFA) comprising an overexpressed gene selected from the group consisting of s fabH2, BKD (lpdV-bkdAA-bkd-AB-bkdB), lplA, fadR, phaC, phaG, alkK, acc, alsS-ilvCD, and leuAmodBCD.
5 . The non-naturally occurring Pseudomonas sp. of claim 4 wherein the Pseudomonas sp. is P. putida KT 2440.
6 . The non-naturally occurring Pseudomonas sp. of claim 4 wherein the gene is s fabH2.
7 . The non-naturally occurring Pseudomonas sp. of claim 4 wherein the gene is BKD (lpdV-bkdAA-bkd-AB-bkdB).
8 . The non-naturally occurring Pseudomonas sp. of claim 4 wherein the gene is lplA.
9 . The non-naturally occurring Pseudomonas sp. of claim 4 wherein the gene is fadR.
10 . The non-naturally occurring Pseudomonas sp. of claim 4 wherein the gene is phaC.
11 . The non-naturally occurring Pseudomonas sp. of claim 4 wherein the gene is phaG.
12 . The non-naturally occurring Pseudomonas sp. of claim 4 wherein the gene is alkK.
13 . The non-naturally occurring Pseudomonas sp. of claim 4 wherein the gene is acc.
14 . The non-naturally occurring Pseudomonas sp. of claim 4 wherein the gene is alsS-ilvCD.
15 . The non-naturally occurring Pseudomonas sp. of claim 4 wherein the gene is leuAmodBCD.