Abstract
Shikimic acid (SA) is an intermediate of the SA pathway that is present in bacteria and plants. SA has gained great interest because it is a precursor in the synthesis of the drug oseltamivir phosphate (OSF), an efficient inhibitor of the neuraminidase enzyme of diverse seasonal influenza viruses, the avian influenza virus H5N1, and the human influenza virus H1N1. For the purposes of OSF production, SA is extracted from the pods of Chinese star anise plants (Illicium spp.), yielding up to 17% of SA (dry basis content). The high demand for OSF necessary to manage a major influenza outbreak is not adequately met by industrial production using SA from plants sources. As the SA pathway is present in the model bacteria Escherichia coli, several “intuitive” metabolically engineered strains have been applied for its successful overproduction by biotechnological processes, resulting in strains producing up to 71 g/L of SA, with high conversion yields of up to 0.42 (mol SA/mol Glc), in both batch and fed-batch cultures using complex fermentation broths, including glucose as a carbon source and yeast extract. Global transcriptomic analyses have been performed in SA-producing strains, resulting in the identification of possible key target genes for the design of a rational strain improvement strategy. Because possible target genes are involved in the transport, catabolism, and interconversion of different carbon sources and metabolic intermediates outside the central carbon metabolism and SA pathways, as genes involved in diverse cellular stress responses, the development of rational cellular strain improvement strategies based on omics data constitutes a challenging task to improve SA production in currently overproducing engineered strains. In this review, we discuss the main metabolic engineering strategies that have been applied for the development of efficient SA-producing strains, as the perspective of omics analysis has focused on further strain improvement for the production of this valuable aromatic intermediate.
Highlights
Compounds derived from the aromatic amino acid (AA) pathway play important roles in the pharmaceutical and food industries as raw materials, additives, or final products (Patnaik et al, 1995; Bongaerts, 2001; Báez et al, 2001; Yi et al, 2002; Chandran et al, 2003; Báez-Viveros et al, 2004; Gosset, 2009)
shikimic acid (SA) is a key intermediate of the common aromatic pathway with diverse applications in the synthesis of valuable pharmaceutical compounds, but major interest relies on SA as the precursor for the chemical synthesis of OSF, the neuraminidase inhibitor of diverse influenza viruses, including pandemic strains
Diverse efforts have been made to produce high titers and yields of SA in metabolically engineered strains of E. coli with successful genetic modifications, including the following: (1) interruption of the SA pathway by the inactivation of shikimate kinase coding genes, which results in the high accumulation of SA; (2) increasing the intracellular availability of the central carbon metabolism (CCM) intermediate PEP by inactivation of the phosphotransferase system (PTS) system and replacing this glucose translocation system by other housekeeping or heterologous glucose transporters and by inactivation of the pykF gene; and (3) the overexpression of diverse key genes of the CCM and SA pathways, such as zwf, tktA, aroB, aroD, and aroE, under the control of constitutively expressed or inducible promoters in plasmid-cloned operons or chromosome-integrated copies
Summary
Compounds derived from the aromatic amino acid (AA) pathway play important roles in the pharmaceutical and food industries as raw materials, additives, or final products (Patnaik et al, 1995; Bongaerts, 2001; Báez et al, 2001; Yi et al, 2002; Chandran et al, 2003; Báez-Viveros et al, 2004; Gosset, 2009). SA has great pharmaceutical relevance because it is the precursor for the chemical synthesis of oseltamivir phosphate (OSF), known as Tamiflu®, used as the antiviral inhibitor of the neuraminidase enzyme for the treatment of diverse seasonal influenza viruses, including influenza A and B, the avian influenza virus H5N1, and the human influenza virus H1N1 (Krämer et al, 2003; Estevez and Estevez, 2012; Ghosh et al, 2012; Diaz Quiroz et al, 2014) For this purpose, SA is obtained from the seed of the Chinese star anise plant Illicium verum, which contains between 2 and 7% of the intermediate. The goal of this work is to review the literature on the great biotechnological achievements made for SA production, mainly in Escherichia coli, and to outline future perspectives on research performed in the omics era, which could provide relevant tools for understanding cell behavior and production optimization via biotechnological processes
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