Accumulation Of Saturated Fatty Acids Research Articles

Inactivation of the inhA-encoded fatty acid synthase II (FASII) enoyl-acyl carrier protein reductase induces accumulation of the FASI end products and cell lysis of Mycobacterium smegmatis.

The mechanism of action of isoniazid (INH), a first-line antituberculosis drug, is complex, as mutations in at least five different genes (katG, inhA, ahpC, kasA, and ndh) have been found to correlate with isoniazid resistance. Despite this complexity, a preponderance of evidence implicates inhA, which codes for an enoyl-acyl carrier protein reductase of the fatty acid synthase II (FASII), as the primary target of INH. However, INH treatment of Mycobacterium tuberculosis causes the accumulation of hexacosanoic acid (C(26:0)), a result unexpected for the blocking of an enoyl-reductase. To test whether inactivation of InhA is identical to INH treatment of mycobacteria, we isolated a temperature-sensitive mutation in the inhA gene of Mycobacterium smegmatis that rendered InhA inactive at 42 degrees C. Thermal inactivation of InhA in M. smegmatis resulted in the inhibition of mycolic acid biosynthesis, a decrease in hexadecanoic acid (C(16:0)) and a concomitant increase of tetracosanoic acid (C(24:0)) in a manner equivalent to that seen in INH-treated cells. Similarly, INH treatment of Mycobacterium bovis BCG caused an inhibition of mycolic acid biosynthesis, a decrease in C(16:0), and a concomitant accumulation of C(26:0). Moreover, the InhA-inactivated cells, like INH-treated cells, underwent a drastic morphological change, leading to cell lysis. These data show that InhA inactivation, alone, is sufficient to induce the accumulation of saturated fatty acids, cell wall alterations, and cell lysis and are consistent with InhA being a primary target of INH.

Saccharomyces cerevisiae cell fatty acid composition and release during fermentation without aeration and in absence of exogenous lipids

Medium-chain fatty-acids (MCFA) are among the main aroma compounds of fermented beverages. High concentrations of MCFA have been found in sluggish and stuck fermentations. It has been suggested that they arrest cell growth, as they may be toxic, but the causes of sluggish and stuck fermentations are still unclear. The aim of this work was to see whether the production of MCFA is related to fatty acid synthesis in the absence of exogenous lipids and aeration, and whether their increase can be regarded as a consequence, instead of the cause, of sluggish and stuck fermentations. Two possibilities were considered: (i) MCFA are produced to replace unsaturated fatty acids (UFA) for cell membrane fluidity when the lack of oxygen makes desaturation of saturated fatty acids (SFA) impossible; or (ii) MCFA are produced following the release of medium-chain acyl-CoA from the fatty acid synthetase complex (FAS) due to the accumulation of SFA, and their hydrolysis to recycle CoA-SH. In the first hypothesis, MCFA should be active in cell metabolism and be found in cell structures; in the second, MCFA should be a discard and prevalently found outside the cell. We carried out a Saccharomyces cerevisiae fermentation experiment in a synthetic, lipid-free medium without aeration. We measured the fatty acid composition of yeast cells and the amounts of MCFA and their ethyl esters in the medium throughout the fermentation. Cell growth and the oxygen content of the medium were also monitored. We found that MCFA are not immobilized in cell structures, but mainly released into the medium. Cell growth is arrested because fatty acid biosynthesis is prevented by the lack of oxygen. The higher MCFA concentrations found in sluggish and stuck fermentations can be thus regarded as an effect, and not the cause, of this arrest. Some suggestions for the prevention of these events are proposed.

Accumulation of unsaturated acylcarnitine molecular species during acute myocardial ischemia: metabolic compartmentalization of products of fatty acyl chain elongation in the acylcarnitine pool.

Long-chain acylcarnitines accumulate during myocardial ischemia and contribute to membrane dysfunction in ischemic zones. On the basis of the 3-fold selectivity for saturated fatty acid accumulation during myocardial ischemia, it was implicitly assumed that saturated long chain acylcarnitine molecular species predominantly accumulated in ischemic myocardium. By exploiting the analytical power of electrospray ionization mass spectroscopy, we now report that unsaturated acylcarnitines are the predominant molecular species of acylcarnitine which accumulate during myocardial ischemia (rank order: octadecadienoyl carnitine > octadecanoyl carnitine > hexadecanoyl carnitine > octadecanoyl carnitine). The aliphatic chain distribution of myocardial acylcarnitine molecular species identified by electrospray ionization mass spectroscopy was independently substantiated by sequential HPLC purification and capillary gas chromatography. Detailed analysis of the individual molecular species of long-chain acylcarnitine demonstrated that fatty acyl chain elongation was prominent in ischemic myocardium (e.g., following 20 min of ischemia, greater than 15% of the accumulated acylcarnitines consisted of 20-carbon unsaturated molecular species). Chain-elongated lipids were essentially confined to the long chain acylcarnitine pool since [9,10-3H]octadec-9'-enoic acid was converted to [3H]eicosenoyl carnitine (12% of the radiolabeled acylcarnitine pool) in ischemic hearts without substantive amounts of [3H]eicosenoyl residues in the fatty acid, triglyceride, and phospholipid pools. Collectively, these results demonstrate the preponderance of unsaturated acylcarnitines in ischemic myocardium and document the metabolic compartmentation of downstream products of fatty acyl chain elongation in the acylcarnitine pool during ischemia.

Evidence for the involvement of unsaturated fatty acids in initiating chromosome replication in Escherichia coli

The analogue 3-decynoyl- N-acetylcysteamine inhibits the synthesis of unsaturated fatty acids in Escherichia coli, resulting in the accumulation of saturated fatty acids in the membrane (Kass, 1968). In the presence of this analogue, DNA, RNA and protein synthesis continue at a linear rate for approximately two doubling times, and then cease. On the other hand, the analogue will inhibit the formation of new replication forks (premature initiation), which normally arise as a result of thymine starvation. Unlike other temperature-sensitive DNA mutants, mutants that are defective in initiating DNA replication ( dnaA or dnaC) are unable to replicate DNA at a permissive temperature if they terminate replication at 42 °C in the presence of 3-decynoyl- N-acetylcysteamine. When replication is terminated at 42 °C, cultures of dnaA or dnaC mutants normally will reinitiate replication upon lowering the temperature to 30 °C. For each mutant this reinitiation is characterized by a particular temperature sensitivity. Such mutants become more temperature sensitive if the temperature is lowered in the presence of 3-decynoyl- N-acetylcysteamine. All the effects of this analogue can be reversed by the addition of unsaturated fatty acids. These results are interpreted using a model in which replication is initiated at a particular lipid site on the membrane. In the absence of unsaturated fatty acids functional lipid sites are not made. Functional sites, however, can be used again provided they are not inactivated by interaction with an inactive dnaA or dnaC product.