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The biotechnological production of dicarboxylic acids (C4) from renewable carbon sources represents an attractive approach for the provision of these valuable compounds by green chemistry means. Glycerol has become a waste product of the biodiesel industry that serves as a highly reduced carbon source for some microorganisms. Escherichia coli is capable of consuming glycerol to produce succinate under anaerobic fermentation, but with the deletion of some tricarboxylic acid (TCA) cycle genes, it is also able to produce succinate and malate in aerobiosis. In this study, we investigate possible rate-limiting enzymes by overexpressing the C-feeding anaplerotic enzymes Ppc, MaeA, MaeB, and Pck in a mutant that lacks the succinate dehydrogenase (Sdh) enzyme. The overexpression of the TCA enzyme Mdh and the activation of the glyoxylate shunt was also examined. Using this unbiased approach, we found that phosphoenol pyruvate carboxylase (Ppc) overexpression enhances an oxidative pathway that leads to increasing succinate, while phosphoenol pyruvate carboxykinase (Pck) favors a more efficient reductive branch that produces mainly malate, at 57.5% of the theoretical maximum molar yield. The optimization of the culture medium revealed the importance of bicarbonate and pH in the production of malate. An additional mutation of the ppc gene highlights its central role in growth and C4 production.

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Biological desulfurization has proven to be a process that is technically and economically feasible on using biotrickling filters that can be performed under aerobic and anoxic conditions. However, microbial communities are different mainly due to the use of different final electron acceptors. The analysis of microbial communities in these systems has not been addressed with regard to the anoxic process. The aim of the work reported here was to analyse the eubacterial community in the two types of bioreactor along the packed bed and during the operation time. The analysis was carried out using the 16S PCR-DGGE molecular fingerprint technique. The microbial profile analysis in the aerobic bioreactor revealed that the community was more diverse and stratified compared to those obtained in the two anoxic bioreactors, influenced by environmental factors. The main OTU involved in this process is genus Thiobacillus, although different species were detected depending on each operational condition.