Significance: Microorganisms in the gastrointestinal tract interact with their host in many ways. Lipid metabolism by gastrointestinal microbes generates multiple fatty acid species that can affect host health. In the representative gut bacterium Lactobacillus plantarum , we revealed a fatty acid metabolism, saturation metabolism of polyunsaturated fatty acid, that generates hydroxy fatty acids, oxo fatty acids, conjugated fatty acids, and partially saturated trans -fatty acids as intermediates. Furthermore, fatty acid analysis in mice suggests that the fatty acid metabolism by gastrointestinal microbes modifies fatty acid composition of the host. Therefore, functional investigations of lipid metabolisms of gastrointestinal microbes may provide new methods for improving our health by altering lipid metabolism related to the onset of metabolic syndrome.

Gut microbial metabolites of polyunsaturated fatty acids have attracted much attention because of their various physiological properties. Dysfunction of tight junction (TJ) in the intestine contributes to the pathogenesis of many disorders such as inflammatory bowel disease. We evaluated the effects of five novel gut microbial metabolites on tumor necrosis factor (TNF)-α-induced barrier impairment in Caco-2 cells and dextran sulfate sodium-induced colitis in mice. 10-Hydroxy-cis-12-octadecenoic acid (HYA), a gut microbial metabolite of linoleic acid, suppressed TNF-α and dextran sulfate sodium-induced changes in the expression of TJ-related molecules, occludin, zonula occludens-1, and myosin light chain kinase. HYA also suppressed the expression of TNF receptor 2 (TNFR2) mRNA and protein expression in Caco-2 cells and colonic tissue. In addition, HYA suppressed the protein expression of TNFR2 in murine intestinal epithelial cells. Furthermore, HYA significantly up-regulated G protein-coupled receptor (GPR) 40 expression in Caco-2 cells. It also induced [Ca²⁺]_i responses in HEK293 cells expressing human GPR40 with higher sensitivity than linoleic acid, its metabolic precursor. The barrier-recovering effects of HYA were abrogated by a GPR40 antagonist and MEK inhibitor in Caco-2 cells. Conversely, 10-hydroxyoctadacanoic acid, which is a gut microbial metabolite of oleic acid and lacks a carbon-carbon double bond at Δ12 position, did not show these TJ-restoring activities and down-regulated GPR40 expression. Therefore, HYA modulates TNFR2 expression, at least partially, via the GPR40-MEK-ERK pathway and may be useful in the treatment of TJ-related disorders such as inflammatory bowel disease.

The filamentous fungus Mortierella alpina 1S-4 is capable of accumulating a large amount of triacylglycerol containing C20 polyunsaturated fatty acids (PUFAs). Indeed, triacylglycerol production by M. alpina 1S-4 can reach 20 g/L of culture broth, and the critical cellular signaling and structural PUFA arachidonic acid (ARA) comprises 30%-70% of the total fatty acid. The demonstrated health benefits of functional PUFAs have in turn encouraged the search for rich sources of these compounds, including fungal strains showing enhanced production of specific PUFAs. Screening for mutants and targeted gene manipulation of M. alpina 1S-4 have elucidated the functions of various enzymes involved in PUFA biosynthesis and established lines with improved PUFA productivity. In some cases, these strains have been used for indistrial-scale production of PUFAs, including ARA. In this review, we described practical ARA production through mutant breeding, functional analyses of genes encoding enzymes involved in PUFA biosynthesis, and recent advances in the production of specific PUFAs through molecular breeding of M. alpina 1S-4.

Hydroxy FAs, one of the gut microbial metabolites of PUFAs, have attracted much attention because of their various bioactivities. The purpose of this study was to identify lactic acid bacteria with the ability to convert linoleic acid to hydroxy FAs. A screening process revealed that a gut bacterium Lactobacillus acidophilus NTV001 converts linoleic acid mainly into 13-hydroxy-cis-9-octadecenoic acid, and resulted in the identification of the hydratase responsible, FA-HY1. Recombinant FA-HY1 was purified and its enzymatic characteristics were investigated. FA-HY1 could convert not only C18 PUFAs but also C20 and C22 PUFAs. C18 PUFAs with a cis carbon-carbon double bond at the Δ12 position were converted into the corresponding 13-hydroxy FAs. Arachidonic acid and DHA were converted into the corresponding 15-hydroxy FA and 14-hydroxy FA, respectively. To the best of our knowledge, this is the first report of a bacterial FA hydratase that can convert C20 and C22 PUFAs into the corresponding hydroxy FAs. These novel hydroxy FAs produced by using FA-HY1 should contribute to elucidate the bioactivities of hydroxy FAs.

We determined the enzymatic characteristics of an industrially important biocatalyst, α-ketoglutarate-dependent l -isoleucine dioxygenase (IDO), which was found to be the enzyme responsible for the generation of (2 S ,3 R ,4 S )-4-hydroxyisoleucine in Bacillus thuringiensis 2e2. Depending on the amino acid used as the substrate, IDO catalyzed three different types of oxidation reactions: hydroxylation, dehydrogenation, and sulfoxidation. IDO stereoselectively hydroxylated several hydrophobic aliphatic l -amino acids, as well as l -isoleucine, and produced ( S )-3-hydroxy- l - allo -isoleucine, 4-hydroxy- l -leucine, ( S )-4-hydroxy- l -norvaline, 4-hydroxy- l -norleucine, and 5-hydroxy- l -norleucine. The IDO reaction product of l -isoleucine, (2 S ,3 R ,4 S )-4-hydroxyisoleucine, was again reacted with IDO and dehydrogenated into (2 S ,3 R )-2-amino-3-methyl-4-ketopentanoate, which is also a metabolite found in B. thuringiensis 2e2. Interestingly, IDO catalyzed the sulfoxidation of some sulfur-containing l -amino acids and generated l -methionine sulfoxide and l -ethionine sulfoxide. Consequently, the effective production of various modified amino acids would be possible using IDO as the biocatalyst.

Linoleic acid Δ9 hydratase, which is involved in linoleic acid saturation metabolism of Lactobacillus plantarum AKU 1009a, was cloned, expressed as a his-tagged recombinant enzyme, purified with an affinity column, and characterized. The enzyme required FAD as a cofactor and its activity was enhanced by NADH. The maximal activities for the hydration of linoleic acid and for the dehydration of 10-hydroxy-cis-12-octadecenoic acid (HYA) were observed at 37 °C in buffer at pH 5.5 containing 0.5 M NaCl. Free C16 and C18 fatty acids with cis-9 double bonds and 10-hydroxy fatty acids served as substrates for the hydration and dehydration reactions, respectively. The apparent Km value for linoleic acid was estimated to be 92 μM, with a kcat of 2.6∙10(-2) s(-1) and a Hill factor of 3.3. The apparent Km value for HYA was estimated to be 98 μM, with a kcat of 1.2∙10(-3) s(-1).