A bacterial biosynthetic pathway for methylated furan fatty acids

Rachelle A.S Lemke,Stephanie M Olson,Kaitlin Morse,Steven D Karlen,Alan Higbee,Emily T Beebe,John Ralph,Joshua J Coon,Brian G Fox,Timothy J Donohue

doi:10.1074/jbc.ra120.013697

Abstract

Fatty acids play many important roles in cells and also in industrial processes. Furan fatty acids (FuFAs) are present in the lipids of some plant, fish, and microbial species and appear to function as second messengers in pathways that protect cells from membrane-damaging agents. We report here the results of chemical, genetic, and synthetic biology experiments to decipher the biosynthesis of the monomethylated FuFA, methyl 9-(3-methyl-5-pentylfuran-2-yl) nonanoate (9M5-FuFA), and its dimethyl counterpart, methyl 9-(3,4-dimethyl-5-pentylfuran-2-yl) nonanoate (9D5-FuFA), in two α-proteobacteria. Each of the steps in FuFA biosynthesis occurs on pre-existing phospholipid fatty acid chains, and we identified pathway intermediates and the gene products that catalyze 9M5-FuFA and 9D5-FuFA synthesis in Rhodobacter sphaeroides 2.4.1 and Rhodopseudomonas palustris CGA009. One previously unknown pathway intermediate was a methylated diunsaturated fatty acid, (10E,12E)-11-methyloctadeca-10,12-dienoic acid (11Me-10t,12t-18:2), produced from (11E)-methyloctadeca-11-enoic acid (11Me-12t-18:1) by a newly identified fatty acid desaturase, UfaD. We also show that molecular oxygen (O2) is the source of the oxygen atom in the furan ring of 9M5-FuFA, and our findings predict that an O2-derived oxygen atom is incorporated into 9M5-FuFA via a protein, UfaO, that uses the 11Me-10t,12t-18:2 fatty acid phospholipid chain as a substrate. We discovered that R. palustris also contains a SAM-dependent methylase, FufM, that produces 9D5-FuFA from 9M5-FuFA. These results uncover the biochemical sequence of intermediates in a bacterial pathway for 9M5-FuFA and 9D5-FuFA biosynthesis and suggest the existence of homologs of the enzymes identified here that could function in FuFA biosynthesis in other organisms.

Highlights

Fatty acids play many important roles in cells and in industrial processes
Fatty acid methyl esters (FAMEs) prepared from the phospholipids found in R. sphaeroides DchrR cells contain higher detectable levels of 9M5-furan fatty acid (FuFA) and 11Me-12t-18:1 than WT cells, in addition to the 18:1, 18:0, 16:1, and 16:0 fatty acid chains found in WT cells (Fig. 2A)
This work reports new information on the biosynthetic pathway and intermediates in the synthesis of two FuFAs (9M5-FuFA and 9D5-FuFA) and gene products (UfaD, UfaO, and FufM) involved in the production of these FuFAs. Both mono- and dimethyl-FuFAs are synthesized from 18:1 fatty acyl chains on phospholipids, and 9D5-FuFA is synthesized

Summary

Introduction

Furan fatty acids (FuFAs) are present in the lipids of some plant, fish, and microbial species and appear to function as second messengers in pathways that protect cells from membrane-damaging agents. We discovered that R. palustris contains a SAM-dependent methylase, FufM, that produces 9D5-FuFA from 9M5-FuFA These results uncover the biochemical sequence of intermediates in a bacterial pathway for 9M5-FuFA and 9D5-FuFA biosynthesis and suggest the existence of homologs of the enzymes identified here that could function in FuFA biosynthesis in other organisms. FuFAs containing zero, one, or two methyl groups are implicated as second messengers in pathways that protect cells from the toxic effects of membrane-damaging agents [15,16,17]. Strategies to produce large quantities of natural or modified FuFAs are limited by a lack of information about their biosynthesis

Results

Discussion

Conclusion