Linkage of Marine Bacterial Polyunsaturated Fatty Acid and Long-Chain Hydrocarbon Biosynthesis.

Marco N. Allemann,Eric E. Allen,Christine N. Shulse

doi:10.3389/fmicb.2019.00702

Marco N. Allemann, Eric E. Allen + Show 1 more

Open Access

https://doi.org/10.3389/fmicb.2019.00702

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Abstract

Various marine gamma-proteobacteria produce omega-3 polyunsaturated fatty acids, such as eicosapentaenoic acid (20:5, EPA) and docosahexaenoic acid (22:6, DHA), which are incorporated into membrane phospholipids. Five genes, designated pfaABCDE, encode the polyketide/fatty acid synthase necessary for production of these long-chain fatty acids. In addition to de novo biosynthesis of EPA and DHA, the “Pfa synthase” is also involved with production of a long-chain polyunsaturated hydrocarbon product (31:9, PUHC) in conjunction with the oleABCD hydrocarbon biosynthesis pathway. In this work, we demonstrate that OleA mediates the linkage between these two pathways in vivo. Co-expression of pfaA-E along with oleA from Shewanella pealeana in Escherichia coli yielded the expected product, a 31:8 ketone along with a dramatic ∼10-fold reduction in EPA content. The decrease in EPA content was independent of 31:8 ketone production as co-expression of an OleA active site mutant also led to identical decreases in EPA content. We also demonstrate that a gene linked with either pfa and/or ole operons in diverse bacterial lineages, herein designated pfaT, plays a role in maintaining optimal production of Pfa synthase derived products in Photobacterium and Shewanella species.

Highlights

Our results indicate that OleA can interact with the Pfa synthase directly, most likely with the acyl carrier protein (ACP) domains that shuttle acyl intermediates, including the 16:4n-3 PUHC precursor molecules, among catalytic domains during de novo polyunsaturated fatty acids (PUFAs) biosynthesis
Given the relationship between the two pathways, it was hypothesized that previously observed differences in EPA content amongst members of the Shewanella genus (Kato and Nogi, 2001), may be due to the diversion of acyl substrates from the EPA pathway into the PUHC pathway
In this work we have demonstrated that bacterial PUFA and PUHC biosynthesis are linked and this linkage is mediated by OleA

Summary

Introduction

The pathway for biosynthesis of omega-3 polyunsaturated fatty acids (PUFAs) in certain marine bacteria occurs via a polyketide synthase type mechanism encoded by five genes pfaABCDE (Metz et al, 2001; Allen and Bartlett, 2002; Shulse and Allen, 2011; Yoshida et al, 2016; Allemann and Allen, 2018). The biosynthesis of long-chain fatty acid-derived olefin hydrocarbons has been previously shown to be the result of the oleABCD pathway in various marine and non-marine bacterial lineages (Beller et al, 2010; Sukovich et al, 2010a,b). In this pathway, hydrocarbons are generated by the condensation of two fatty acyl chains via a “head-to-head” Claisen condensation mechanism, resulting in an intermediate β-keto acid product (Figure 1; Frias et al, 2011; Goblirsch et al, 2016). In the absence of downstream processing by OleBCD, the OleA catalyzed β-keto acid intermediate undergoes spontaneous decarboxylation to form a ketone product (Sukovich et al, 2010b; Frias et al, 2011; Figure 1)

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