The idea of renewable and regenerative resources has inspired research for more than a hundred years. Ideally, the only spent energy will replenish itself, like plant material, sunlight, thermal energy or wind. Biodiesel or ethanol are examples, since their production relies mainly on plant material. However, it has become apparent that crop derived biofuels will not be sufficient to satisfy future energy demands. Thus, especially in the last decade a lot of research has focused on the production of next generation biofuels. A major subject of these investigations has been the microbial fatty acid biosynthesis with the aim to produce fatty acids or derivatives for substitution of diesel. As an industrially important organism and with the best studied microbial fatty acid biosynthesis, Escherichia coli has been chosen as producer in many of these studies and several reviews have been published in the fields of E. coli fatty acid biosynthesis or biofuels. However, most reviews discuss only one of these topics in detail, despite the fact, that a profound understanding of the involved enzymes and their regulation is necessary for efficient genetic engineering of the entire pathway. The first part of this review aims at summarizing the knowledge about fatty acid biosynthesis of E. coli and its regulation, and it provides the connection towards the production of fatty acids and related biofuels. The second part gives an overview about the achievements by genetic engineering of the fatty acid biosynthesis towards the production of next generation biofuels. Finally, the actual importance and potential of fatty acid-based biofuels will be discussed.

This study investigated the production of triacylglycerols in cells of the wild type of Escherichia coli and of a strain with a deleted diacylglycerol kinase gene (dgkA). By overexpression of atfA from Acinetobacter baylyi ADP1 and fadD from E. coli in the dgkA deletion mutant, cellular contents of up to 4.9% (w/w) triacylglycerols could be achieved in batch cultivation. Furthermore, heterologous expression of atfA relieves the negative effects of dgkA deletion on growth. Process optimization and fed-batch fermentation resulted in the production of 530 mg l (-1) triacylglycerols and a maximal content of 8.5% (w/w) triacylglycerols of the cell dry mass. This clearly exceeded all previous results concerning triacylglycerol production in E. coli. Furthermore, the production of extracellular free fatty acids and fatty acid ethyl esters was investigated. Like triacylglycerols, both products are potential biofuels, and we show their continuous production in a repeated batch process, with recovery of the production cells.

The refinement of biodiesel or renewable diesel from bacterial lipids has a great potential to make a contribution for energy production in the future. This study provides new data concerning suitable nutrient concentrations for cultivation of the Gram-positive Rhodococcus opacus PD630, which is able to accumulate large amounts of lipids during nitrogen limitation. Enhanced concentrations of magnesium have been shown to increase the final optical density and the lipid content of the cells. Elevated phosphate concentrations slowed down the onset of the accumulation phase, without a clear effect on the final optical density and the cell’s lipid content. A robust growth of R. opacus was possible in the presence of ammonium concentrations of up to 1.4 g l-1 and sucrose concentrations of up to 240 g l-1, with an optimum regarding growth and lipid storage observed in the range of 0.2 to 0.4 g l-1 ammonium and 20 to 40 g l-1 sucrose, respectively. Moreover, R. opacus showed tolerance to high salt concentrations.