Unexpected contribution of the Fak system and the thioesterase TesE to the growth and membrane physiology of <i>Enterococcus faecalis</i>

<i>Enterococcus faecalis</i> is a native of the intestine and a hospital-acquired pathogen that uses host fatty acids to form its membrane. We investigated the utilization of exogenous fatty acids via the fatty acid kinase (Fak) system to understand the varied impacts fatty acids have on physiology. FakB proteins bind fatty acids, and FakA then phosphorylates them for lipid synthesis. Network analysis indicated that two of the four FakB proteins of <i>E. faecalis</i> OG1RF cluster with described proteins of <i>Staphylococcus aureus</i> and <i>Streptococcus pneumoniae</i> (FakB1, FakB2). However, two additional <i>E. faecalis</i> FakB proteins clustered separately and distinctly from characterized proteins; these were subsequently denoted as FakB4 and FakB5. A strain deleted for three of the four genes (<i>ΔfakB1,2,5</i> strain) had severe morphological defects when grown in rich media. Deletion of all four <i>fakB</i>-encoding genes was not possible unless a thioesterase encoding gene, <i>tesE</i>, was also deleted (<i>Δquint</i> strain). The <i>Δquint</i> strain behaved similarly to wild-type OG1RF in rich media, indicating that the combination of free fatty acids from the growth environment and those liberated via TesE was detrimental to the <i>ΔfakB1,2,5</i> strain. The <i>Δquint</i> strain grew unimpeded in saturated fatty acids that are normally toxic to <i>E. faecalis,</i> indicating that incorporation of these fatty acids into phospholipids mediates their toxicity. While saturated fatty acids reduced the membrane fluidity of wild-type OG1RF, they had no impact on the <i>Δquint</i> strain. Our combined data support that the Fak system in <i>E. faecalis</i> plays a critical role in maintaining membrane fluidity and driving enterococcal physiology.IMPORTANCEBacteria living within humans encounter a variety of fatty acids that they can use to synthesize their own cellular material. However, different fatty acids can have a variety of effects on the same bacterial species. Within, we examined how <i>Enterococcus faecalis</i>, which naturally lives in human intestines but can also cause disease, uses fatty acids from its environment. We discovered unexpectedly that fatty acid binding proteins contribute to many aspects controlling bacterial growth, shape, and behavior.