Abstract
Lactobacilli convert linoleic acid to the antifungal compound 10-hydroxy-12-octadecenoic acid (10-HOE) by linoleate 10-hydratase (10-LAH). However, the effect of this conversion on cellular membrane physiology and properties of the cell surface have not been demonstrated. Moreover, Lactobacillus plantarum produces 13-hydroxy-9-octadecenoic acid (13-HOE) in addition to 10-HOE, but the antifungal activity of 13-HOE was unknown. Phylogenetic analyses conducted in this study did not differentiate between 10-LAH and linoleate 13-hydratase (13-LAH). Thus, linoleate hydratases (LAHs) must be characterized through their differences in their activities of linoleate conversion. Four genes encoding putative LAHs from lactobacilli were cloned, heterologous expressed, purified and identified as FAD-dependent 10-LAH. The unsaturated fatty acid substrates stimulated the growth of lactobacilli. We also investigated the role of 10-LAH in ethanol tolerance, membrane fluidity and hydrophobicity of cell surfaces in lactobacilli by disruption of lah. Compared with the L. plantarum lah deficient strain, 10-LAH in wild-type strain did not exert effect on cell survival and membrane fluidity under ethanol stress, but influenced the cell surface hydrophobicity. Moreover, deletion of 10-LAH in L. plantarum facilitated purification of 13-HOE and demonstration of its antifungal activity against Penicillium roqueforti and Aspergillus niger.
Highlights
Antifungal metabolites of lactic acid bacteria have potential for applications as antifungal preservatives in cereal products, and in silage (Magnusson et al, 2003; Oliveira et al, 2014)
L. plantarum TMW1.460 lah produced 13-hydroxy9-octadecenoic acid (13-HOE) but not 10-HOE or 10,13dihydroxy octadecanoic acid, demonstrating that their formation by L. plantarum TMW1.460 is attributable to a dedicated linoleate 13-hydratase acting on linoleic acid and 10-HOE, respectively
The comparison of products formed by L. plantarum TMS1.460, the 10-linoleate hydratases (LAHs) deficient mutant of this strain, and the 10-LAH of this strain strongly suggest that 13-HOE and 10,13 dihydroxy octadecanoic acid formation by this strain is attributable to a linoleate 13-hydratase that was recently characterized in L. acidophilus (Park et al, 2015)
Summary
Antifungal metabolites of lactic acid bacteria have potential for applications as antifungal preservatives in cereal products, and in silage (Magnusson et al, 2003; Oliveira et al, 2014). Several hydroxy fatty acids have antifungal activity (Hou and Forman Iii, 2000; Hou, 2008) and antifungal 3-hydroxy fatty acids of C10 to C14 chain lengths are formed by Lactobacillus plantarum MiLAB 14 (Sjögren et al, 2003). The FAD containing LAH in S. pyogenes hydrates the cis-9 and cis double bonds of C16 and C18 fatty acids to produce 10hydroxy and 10,13-dihydroxy fatty acids (Volkov et al, 2010). The crystal structure of the LAH from Lactobacillus acidophilus provided the structural basis for the selective substrate recognition of linoleate 10-hydratase (Volkov et al, 2013). A second LAH in L. acidophilus hydrates the cis-12 double bond to produce 13hydroxy fatty acid (Kim et al, 2015; Park et al, 2015)
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