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Uml2 is a novel CalB-type lipase of Ustilago maydis with phospholipase A activity

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Abstract

CalB of Pseudozyma aphidis (formerly named Candida antarctica) is one of the most widely applied enzymes in industrial biocatalysis. Here, we describe a protein with 66 % sequence identity to CalB, designated Ustilago maydis lipase 2 (Uml2), which was identified as the product of gene um01422 of the corn smut fungus U. maydis. Sequence analysis of Uml2 revealed the presence of a typical lipase catalytic triad, Ser-His-Asp with Ser125 located in a Thr-Xaa-Ser-Xaa-Gly pentapeptide. Deletion of the uml2 gene in U. maydis diminished the ability of cells to hydrolyse fatty acids from tributyrin or Tween 20/80 substrates, thus demonstrating that Uml2 functions as a lipase that may contribute to nutrition of this fungal pathogen. Uml2 was heterologously produced in Pichia pastoris and recombinant N-glycosylated Uml2 protein was purified from the culture medium. Purified Uml2 released short- and long-chain fatty acids from p-nitrophenyl esters and Tween 20/80 substrates. Furthermore, phosphatidylcholine substrates containing long-chain saturated or unsaturated fatty acids were effectively hydrolysed. Both esterase and phospholipase A activity of Uml2 depended on the Ser125 catalytic residue. These results indicate that Uml2, in contrast to CalB, exhibits not only esterase and lipase activity but also phospholipase A activity. Thus, by genome mining, we identified a novel CalB-like lipase with different substrate specificities.

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Acknowledgments

This work was partly financed by the Ministry of Innovation, Science and Research of the German state of North-Rhine Westphalia (MIWF) and the European Regional Development Fund (EFRE). We thank the Ministry of Innovation, Science and Research of North Rhine-Westphalia and Heinrich Heine University Düsseldorf for a scholarship within the CLIB2021-Graduate Cluster Industrial Biotechnology.

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Authors and Affiliations

  1. Institut für Molekulare Mykologie, Heinrich-Heine-Universität, Universitätsstr. 1/26.12, 40225 Düsseldorf, Germany

    Christoph Buerth, Joachim F. Ernst & Denis Tielker

  2. Institut für Molekulare Enzymtechnologie, Heinrich-Heine-Universität, Düsseldorf, Germany

    Filip Kovacic, Susanne Wilhelm & Karl-Erich Jaeger

  3. Institut für Mikrobiologie, Heinrich-Heine-Universität, Düsseldorf, Germany

    Janpeter Stock, Marius Terfrüchte, Michael Feldbrügge & Kerstin Schipper

  4. Institut für Molekulare Mykologie, Heinrich-Heine-Universität, Universitätsstrasse 1/26.12, 40225, Düsseldorf, Germany

    Joachim F. Ernst

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  1. Christoph Buerth
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  2. Filip Kovacic
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Correspondence to Joachim F. Ernst.

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Buerth, C., Kovacic, F., Stock, J. et al. Uml2 is a novel CalB-type lipase of Ustilago maydis with phospholipase A activity. Appl Microbiol Biotechnol 98, 4963–4973 (2014). https://doi.org/10.1007/s00253-013-5493-6

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