title: Cyclopropanation of membrane unsaturated fatty acids is not essential to the acid stress response of Lactococcus lactis subsp. cremoris.
abstract: Cyclopropane fatty acids (CFAs) are synthetized in situ by the transfer of a methylene group from S-adenosyl-L-methionine to a double bond of unsaturated fatty acid chains of membrane phospholipids. This conversion, catalyzed by the Cfa synthase enzyme, occurs in many bacteria and is recognized to play a key role in the adaptation of bacteria in response to a drastic perturbation of the environment. The role of CFAs in the acid tolerance response was investigated in the lactic acid bacterium Lactococcus lactis MG1363. A mutant of the cfa gene was constructed by allelic exchange. The cfa gene encoding the Cfa synthase was cloned and introduced into the mutant to obtain the complemented strain for homologous system studies. Data obtained by gas chromatography (GC) and GC-mass spectrometry (GC-MS) validated that the mutant could not produce CFA. The CFA levels in both the wild-type and complemented strains increased upon their entry to stationary phase, especially with acid-adapted cells or, more surprisingly, with ethanol-adapted cells. The results obtained by performing quantitative reverse transcription-PCR (qRT-PCR) experiments showed that transcription of the cfa gene was highly induced by acidity (by 10-fold with cells grown at pH 5.0) and by ethanol (by 9-fold with cells grown with 6% ethanol) in comparison with that in stationary phase. Cell viability experiments were performed after an acidic shock on the mutant strain, the wild-type strain, and the complemented strain, as a control. The higher viability level of the acid-adapted cells of the three strains after 3 h of shock proved that the cyclopropanation of unsaturated fatty acids is not essential for L. lactis subsp. cremoris survival under acidic conditions. Moreover, fluorescence anisotropy data showed that CFA itself could not maintain the membrane fluidity level, particularly with ethanol-grown cells.
authors.affiliation: Laboratoire Recherche en Vigne et Vin EA 4149, Université de Bourgogne, IUVV, 1 Rue Claude Ladrey, Campus Universitaire Montmuzard, BP27877, F-21078 Dijon, France.
authors.forename: Thi Mai Huong
authors.initials: TM
authors.lastname: To
authors.forename: Cosette
authors.initials: C
authors.lastname: Grandvalet
authors.forename: Raphaëlle
authors.initials: R
authors.lastname: Tourdot-Maréchal
mesh_terms: D000143:Acids / Q000633:toxicity*; D003521:Cyclopropanes / Q000378:metabolism*; D005227:Fatty Acids / Q000378:metabolism*; D005231:Fatty Acids, Unsaturated / Q000378:metabolism*; D008401:Gas Chromatography-Mass Spectrometry; D005816:Genetic Complementation Test; D013294:Lactococcus lactis / Q000737:chemistry / Q000187:drug effects / Q000502:physiology*; D008563:Membrane Lipids / Q000378:metabolism*; D008780:Methyltransferases / Q000235:genetics / Q000378:metabolism; D050296:Microbial Viability / Q000187:drug effects; D050505:Mutant Proteins / Q000235:genetics / Q000378:metabolism; D010743:Phospholipids / Q000378:metabolism; D013312:Stress, Physiological
chemical_list: D000143:Acids; D003521:Cyclopropanes; D005227:Fatty Acids; D005231:Fatty Acids, Unsaturated; D008563:Membrane Lipids; D050505:Mutant Proteins; D010743:Phospholipids; C028775:cyclopropane fatty acids; D008780:Methyltransferases; C019964:cyclopropane synthetase