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Overexpression of a phosphatidic acid phosphatase type 2 leads to an increase in triacylglycerol production in oleaginous Rhodococcus strains

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Abstract

Oleaginous Rhodococcus strains are able to accumulate large amounts of triacylglycerol (TAG). Phosphatidic acid phosphatase (PAP) enzyme catalyzes the dephosphorylation of phosphatidic acid (PA) to yield diacylglycerol (DAG), a key precursor for TAG biosynthesis. Studies to establish its role in lipid metabolism have been mainly focused in eukaryotes but not in bacteria. In this work, we identified and characterized a putative PAP type 2 (PAP2) encoded by the ro00075 gene in Rhodococcus jostii RHA1. Heterologous expression of ro00075 in Escherichia coli resulted in a fourfold increase in PAP activity and twofold in DAG content. The conditional deletion of ro00075 in RHA1 led to a decrease in the content of DAG and TAG, whereas its overexpression in both RHA1 and Rhodococcus opacus PD630 promoted an increase up to 10 to 15 % by cellular dry weight in TAG content. On the other hand, expression of ro00075 in the non-oleaginous strain Rhodococcus fascians F7 promoted an increase in total fatty acid content up to 7 % at the expense of free fatty acid (FFA), DAG, and TAG fractions. Moreover, co-expression of ro00075/atf2 genes resulted in a fourfold increase in total fatty acid content by a further increase of the FFA and TAG fractions. The results of this study suggest that ro00075 encodes for a PAP2 enzyme actively involved in TAG biosynthesis. Overexpression of this gene, as single one or with an atf gene, provides an alternative approach to increase the biosynthesis and accumulation of bacterial oils as a potential source of raw material for biofuel production.

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Acknowledgments

The authors thank W.W. Mohn for providing both R. jostii RHA1 and pTip-QC2 vector. This study was financially supported by the SCyT of the University of Patagonia San Juan Bosco, the Agencia Comodoro Conocimiento (MCR), Oil m&s Company, Project PIP-CONICET Nro. 0764, Project PFIP CHU-25 (COFECyT), and Project PICT2012 Nro. 2031 (ANPCyT), Argentina. Alvarez H.M. is a career investigator and Hernández M.A. is a posdoctoral scholarship holder of the Consejo Nacional de Investigaciones Científicas y Técnicas (CONICET), Argentina.

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Authors and Affiliations

  1. Centro Regional de Investigación y Desarrollo Científico Tecnológico, Facultad de Ciencias Naturales, Universidad Nacional de la Patagonia San Juan Bosco, Ruta Provincial N° 1, Km 4-Ciudad Universitaria, 9000, Comodoro Rivadavia, Chubut, Argentina

    Martín A. Hernández & Héctor M. Alvarez

  2. Instituto de Biología Molecular y Celular de Rosario, Consejo Nacional de Investigaciones Científicas y Técnicas, Facultad de Ciencias Bioquímicas y Farmacéuticas, Universidad Nacional de Rosario, Ocampo y Esmeralda, 2000, Rosario, Santa Fe, Argentina

    Santiago Comba, Ana Arabolaza & Hugo Gramajo

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  1. Martín A. Hernández
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  2. Santiago Comba
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  5. Héctor M. Alvarez
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Correspondence to Héctor M. Alvarez.

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Hernández, M.A., Comba, S., Arabolaza, A. et al. Overexpression of a phosphatidic acid phosphatase type 2 leads to an increase in triacylglycerol production in oleaginous Rhodococcus strains. Appl Microbiol Biotechnol 99, 2191–2207 (2015). https://doi.org/10.1007/s00253-014-6002-2

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