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Reprogramming Hansenula polymorpha for penicillin production: expression of the Penicillium chrysogenum pcl gene

Gidijala, L., van der Klei, I. J., Veenhuis, M. & Kiel, J. A. K. W., Oct-2007, In : Fems Yeast Research. 7, 7, p. 1160-1167 8 p.

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We aim to introduce the penicillin biosynthetic pathway into the methylotrophic yeast Hansenula polymorpha. To allow simultaneous expression of the multiple genes of the penicillin biosynthetic pathway, additional markers were required. To this end, we constructed a novel host-vector system based on methionine auxotrophy and the H. polymorpha MET6 gene, which encodes a putative cystathionine beta-lyase. With this new host-vector system, the Penicillium chrysogenum pcl gene, encoding peroxisomal phenylacetyl-CoA ligase (PCL), was expressed in H. polymorpha. PCL has a potential C-terminal peroxisomal targeting signal type 1 (PTS1). Our data demonstrate that a green fluorescent protein-PCL fusion protein has a dual location in the heterologous host in the cytosol and in peroxisomes. Mutation of the PTS1 of PCL (SKI-COOH) to SKL-COOH restored sorting of the fusion protein to peroxisomes only. Additionally, we demonstrate that peroxisomal PCL-SKL produced in H. polymorpha displays normal enzymatic activities.

Original languageEnglish
Pages (from-to)1160-1167
Number of pages8
JournalFems Yeast Research
Volume7
Issue number7
Publication statusPublished - Oct-2007
Event4th Hansenula Polymorpha Worldwide Network Conference (HPWN) - , Netherlands
Duration: 3-Sep-20065-Sep-2006

Event

4th Hansenula Polymorpha Worldwide Network Conference (HPWN)

03/09/200605/09/2006

Netherlands

Event: Other

    Keywords

  • beta-lactam antibiotics, filamentous fungi, genetic engineering, microbody, PTS1 receptor, ISOPENICILLIN-N-ACYLTRANSFERASE, SACCHAROMYCES-CEREVISIAE, PEROXISOME BIOGENESIS, METHANOL METABOLISM, TARGETING SIGNALS, FILAMENTOUS FUNGI, PTS1 PROTEINS, YEAST, BIOSYNTHESIS, MATRIX

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