The alkane-assimilating yeast Yarrowia lipolytica degrades very efficiently hydrophobic substrates such as n-alkanes, fatty acids, fats and oils for which it has specific metabolic pathways. An overview of the oxidative degradation pathways for alkanes and triglycerides in Y. lipolytica is given, with new insights arising from the recent genome sequencing of this yeast. This includes the interaction of hydrophobic substrates with yeast cells, their uptake and transport, the primary alkane oxidation to the corresponding fatty alcohols and then by different enzymes to fatty acids, and the subsequent degradation in peroxisomal beta-oxidation or storage into lipid bodies. Several enzymes involved in hydrophobic substrate utilisation belong to multigene families, such as lipases/esterases (LIP genes), cytochromes P450 (ALK genes) and peroxisomal acyl-CoA oxidases (POX genes). Examples are presented demonstrating that wild-type and genetically engineered strains of Y. lipolytica can be used for alkane and fatty-acid bioconversion, such as aroma production, for production of SCP and SCO, for citric acid production, in bioremediation, in fine chemistry, for steroid biotransformation, and in food industry. These examples demonstrate distinct advantages of Y. lipolytica for their use in bioconversion reactions of biotechnologically interesting hydrophobic substrates.

The consumers' demand for natural flavour and fragrances rises. To be natural, compounds have to result from the extraction of natural materials and/or to be transformed by natural means such as the use of enzymes or whole cells. Fungi are able to transform some fatty acids into lactones that can thus be natural. Although some parts of this subject have been reviewed several times, the present article proposes to review the different pathways utilised, the metabolic engineering strategies and some current concerns on the reactor application of the transformation including scaling up data. The main enzymatic steps are hydroxylation and β-oxidation in the traditional way, and lactone desaturation or Baeyer-Villiger oxidation. Although the pathway to produce γ-decalactone is rather well known, metabolic engineering strategies may result in significant improvements in the productivity. For the production of other lactones, a key step is the hydroxylation of fatty acids. Beside the biotransformation, increasing the production of the various lactones requires from biotechnologists to solve two main problems which are the toxicity of lactones toward the producing cell and the aeration of the emulsified reactor as the biochemical pathway is very sensitive to the level of available oxygen. The strategies employed to resolve these problems will be presented.

Wild-type Escherichia coli K-12 ferments glucose to a mixture of ethanol and acetic, lactic, formic, and succinic acids. In anoxic chemostat culture at four dilution rates and two different oxidoreduction potentials (ORP), this strain generated a spectrum of products which depended on ORP. Whatever the dilution rate tested, in low reducing conditions (؊100 mV), the production of formate, acetate, ethanol, and lactate was in molar proportions of approximately 2.5:1:1:0.3, and in high reducing conditions (؊320 mV), the production was in molar proportions of 2:0.6:1:2. The modification of metabolic fluxes was due to an ORP effect on the synthesis or stability of some fermentation enzymes; thus, in high reducing conditions, lactate dehydrogenasespecific activity increased by a factor of 3 to 6. Those modifications were concomitant with a threefold decrease in acetyl-coenzyme A (CoA) needed for biomass synthesis and a 0.5-to 5-fold decrease in formate flux. Calculations of carbon and cofactor balances have shown that fermentation was balanced and that extracellular ORP did not modify the oxidoreduction state of cofactors. From this, it was concluded that extracellular ORP could regulate both some specific enzyme activities and the acetyl-CoA needed for biomass synthesis, which modifies metabolic fluxes and ATP yield, leading to variation in biomass synthesis.A wealth of information is available on the response of Escherichia coli cellular metabolism to pH, water activity, or temperature variations, but little is known about the action of extracellular oxidoreduction potentials (ORP) on metabolism, although numerous reactions and regulations are of the oxidoreduction type. Previous studies have shown that substrates with different oxidation states yield a specific product spectrum. Thus, with glucose (oxidation state ϭ 0), the spectrum of main end products (formate:acetate:ethanol:lactate) is equal to 2:1:1:2, with glucitol (oxidation state ϭ Ϫ1), it is equal to 2:1:6:0.5, and with glucuronate (oxidation state ϭ 2), it is equal to 1:5:1:1, with small amounts of succinate also being produced in each case (1). Those metabolic flux modifications are also observed when the nature of external electron acceptors varies (7). In the same way, the NADH/NAD ϩ ratio, which is responsible for regulation of enzymes (8) or genes (16), can be influenced by the oxidation level of the substrate (36) or by the availability and nature of electron acceptors (7). In addition, protein folding and disulfide bond formation are regulated and modified by different oxidoreductase enzymes and oxidoreduction couples (glutathione, thioredoxin) (26). Recently, Taylor and Zhulin in their review have shown that ORP influences or could influence numerous regulations of cell functions controlled via Per-Arnt-Sim (PAS)-containing receptors (signaling modules that monitor changes in light, ORP, oxygen, and the overall energy level of a cell), transducers, and regulators (34). Thus, E. coli senses the medium ORP and swims to a preferred ORP niche by redox...

Some microorganisms can transform methyl ricinoleate into ␥-decalactone, a valuable aroma compound, but yields of the bioconversion are low due to (i) incomplete conversion of ricinoleate (C 18 ) to the C 10 precursor of ␥-decalactone, (ii) accumulation of other lactones (3-hydroxy-␥-decalactone and 2-and 3-decen-4-olide), and (iii) ␥-decalactone reconsumption. We evaluated acyl coenzyme A (acyl-CoA) oxidase activity (encoded by the POX1 through POX5 genes) in Yarrowia lipolytica in lactone accumulation and ␥-decalactone reconsumption in POX mutants. Mutants with no acyl-CoA oxidase activity could not reconsume ␥-decalactone, and mutants with a disruption of pox3, which encodes the short-chain acyl-CoA oxidase, reconsumed it more slowly. 3-Hydroxy-␥-decalactone accumulation during transformation of methyl ricinoleate suggests that, in wild-type strains, ␤-oxidation is controlled by 3-hydroxyacyl-CoA dehydrogenase. In mutants with low acyl-CoA oxidase activity, however, the acyl-CoA oxidase controls the ␤-oxidation flux. We also identified mutant strains that produced 26 times more ␥-decalactone than the wild-type parents.