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University of Groningen New Yeast Expression Platforms Based On University of Groningen New yeast expression platforms based on methylotrophic Hansenula polymorpha and Pichia pastoris and on dimorphic Arxula adeninivorans and Yarrowia lipolytica - A comparison Gellissen, G.; Kunze, G.; Gaillardin, C.; Cregg, J.M; Berardi, E.; Veenhuis, M; van der Klei, I.J. Published in: Fems Yeast Research DOI: 10.1016/j.femsyr.2005.06.004 IMPORTANT NOTE: You are advised to consult the publisher's version (publisher's PDF) if you wish to cite from it. Please check the document version below. Document Version Publisher's PDF, also known as Version of record Publication date: 2005 Link to publication in University of Groningen/UMCG research database Citation for published version (APA): Gellissen, G., Kunze, G., Gaillardin, C., Cregg, J. M., Berardi, E., Veenhuis, M., & van der Klei, I. J. (2005). New yeast expression platforms based on methylotrophic Hansenula polymorpha and Pichia pastoris and on dimorphic Arxula adeninivorans and Yarrowia lipolytica - A comparison. Fems Yeast Research, 5(11), 1079 - 1096. https://doi.org/10.1016/j.femsyr.2005.06.004 Copyright Other than for strictly personal use, it is not permitted to download or to forward/distribute the text or part of it without the consent of the author(s) and/or copyright holder(s), unless the work is under an open content license (like Creative Commons). Take-down policy If you believe that this document breaches copyright please contact us providing details, and we will remove access to the work immediately and investigate your claim. Downloaded from the University of Groningen/UMCG research database (Pure): http://www.rug.nl/research/portal. For technical reasons the number of authors shown on this cover page is limited to 10 maximum. Download date: 25-09-2021 FEMS Yeast Research 5 (2005) 1079–1096 www.fems-microbiology.org MiniReview New yeast expression platforms based on methylotrophic Hansenula polymorpha and Pichia pastoris and on dimorphic Arxula adeninivorans and Yarrowia lipolytica – A comparison Gerd Gellissen a, Gotthard Kunze b,*, Claude Gaillardin c, James M. Cregg d, Enrico Berardi e, Marten Veenhuis f, Ida van der Klei f a PharmedArtis GmbH, Forckenbeckstr. 6, 52074 Aachen, Germany b Institut fu¨r Pflanzengenetik und Kulturpflanzenforschung, Correnstr. 3, 06466 Gatersleben, Germany c Ge´ne´tique Moleculaire et Cellulaire, UMR1238 INAPG-INRA-CNRS, Institut National Agronomique, Paris-Grignon, 78850 Thiverval Grignon, France d Keck Graduate Institute of Applied Sciences, 535 Watson Drive, Claremont, CA 91711, USA e Universita` degli Studi, Via Brecce Bianche, 60131 Ancona, Italy f Eukaryotic Microbiology, RUG, Kerklaan 30, 9751 Haren, The Netherlands Received 25 April 2005; received in revised form 9 June 2005; accepted 9 June 2005 First published online 24 August 2005 Abstract Yeasts combine the ease of genetic manipulation and fermentation of a microbial organism with the capability to secrete and to modify proteins according to a general eukaryotic scheme. Yeasts thus provide attractive platforms for the production of recombi- nant proteins. Here, four important species are presented and compared: the methylotrophic Hansenula polymorpha and Pichia pas- toris, distinguished by an increasingly large track record as industrial platforms, and the dimorphic species Arxula adeninivorans and Yarrrowia lipolytica, not yet established as industrial platforms, but demonstrating promising technological potential, as discussed in this article. Ó 2005 Federation of European Microbiological Societies. Published by Elsevier B.V. All rights reserved. Keywords: Hansenula polymorpha; Pichia pastoris; Arxula adeninivorans; Yarrrowia lipolytica; Yeast expression platforms 1. Introduction respect, yeasts offer considerable advantages over alter- native microbial and eukaryotic cellular systems in pro- The exploitation of recombinant-DNA technology to viding low-cost screening and production systems for engineer expression systems for heterologous protein authentically processed and modified compounds. The production has provided a major task during the last four selected organisms furthermore meet safety prereq- decades. Production procedures had to be developed uisites in that they do not harbour pyrogens, pathogens that employ platforms which meet both the demand or viral inclusions. for efficient mass production and criteria of safety and The initial yeast system developed for heterologous authenticity of the produced compounds [1,2]. In this gene expression was based on the bakerÕs yeast Saccha- romyces cerevisiae. This platform has been successfully * applied to the production of various FDA-approved Corresponding author. Tel.: +49 394 825 247; fax: +49 394 825 366. pharmaceuticals, including insulin [3] and HBsAg [4]. E-mail address: [email protected] (G. Kunze). However, when using this system, certain limitations 1567-1356/$22.00 Ó 2005 Federation of European Microbiological Societies. Published by Elsevier B.V. All rights reserved. doi:10.1016/j.femsyr.2005.06.004 1080 G. Gellissen et al. / FEMS Yeast Research 5 (2005) 1079–1096 and drawbacks are often encountered, since S. cerevisiae recent book on production of recombinant proteins tends to hyperglycosylate recombinant proteins; and various chapters therein [1,2,6,7,9,11]. N-linked carbohydrate chains are terminated by man- Despite the superiority of some characteristics in the nose attached to the chain via an a1,3 bond, which is examples described below, there is clearly no single sys- considered to be allergenic. Other restrictions are the tem that is optimal for the production of all possible consequence of the limited variety of carbon sources proteins. Predictions of a successful development for a that can be utilised by this species, which limits the fer- given protein can only be made to a certain extent when mentation design options. Sometimes, the preferential restricting the initial strain engineering to a single spe- use of episomal vectors leads to instabilities of recombi- cies. The availability of a wide-range yeast vector system nant strains; as a result, batch inconsistencies of produc- enables the assessment of several yeasts in parallel for tion runs can be of major concern [2]. their capability to produce a particular protein in de- Therefore alternative yeast systems have been defined sired amounts and quality [20–22]. Applied to the four that can potentially overcome the described limitations selected, divergent species a high probability of success of the traditional bakerÕs yeast. The following compari- can be envisaged. son includes four platforms, chosen as examples out of a wide range of yeast-based systems now available: the two methylotrophic yeast species Hansenula polymorpha 2. History, phylogenetic position, basic genetics and (H.p.) [5,6] and Pichia pastoris (P.p.) [5,7], and the two biochemistry of the four selected yeast species dimorphic organisms Arxula adeninivorans (A.a.) [8,9] and Yarrowia lipolytica (Y.l.) [10,11]. The selected 2.1. H. polymorpha and P. pastoris organisms share the capability to utilize a broad range of carbon sources, two of them (H.p. and A.a.) can A limited number of yeast species is able to utilize assimilate nitrate, all but P.p. are thermo-tolerant spe- methanol as sole energy and carbon source. These cies with A.a. and Y.l. exhibiting a temperature-depen- include H. polymorpha, P. pastoris, Candida boidinii dent dimorphism with hyphae formed at elevated and P. methanolica [5]. The first methylotrophic yeast temperature. For all selected systems a range of host described was Kloeckera sp. No. 2201, later re-identified strains and relevant genetic elements is available. as C. boidinii [23]. Subsequently other species, including In case of the two methylotrophic species, engineered H. polymorpha and P. pastoris, were identified as having strains have been developed that exhibit human-like methanol-assimilating capabilities [24]. The taxonomic N-glycosylation patterns (H.-A. Kang, pers. commun.) position of the two species is shown in Fig. 1. [12]. The genome of both species has been completely P. pastoris was initially chosen for the production of deciphered (J.M. Cregg, pers. commun.) and for H.p. single-cell proteins (SCP) for feedstock due to its effi- a microarray chip is available [13,14]. Accordingly both cient growth in methanol-containing media, since syn- species are distinguished by an expanding track record thesis of methanol from natural methane was as production hosts for industrial and pharmaceutical inexpensive in the late 60s. However, as the costs of proteins, for both systems efficient secretion of recombi- methane increased in the oil crisis in the 1970s, and that nant proteins (>10 g lÀ1) have been reported [15,16]. All of soy beans, the major feedstock source, decreased, of this attests to the favourable characteristics of these production of SCP in this yeast never became economi- two yeast species. In the case of the two dimorphic spe- cally competitive [25,26]. With the methods of recombi- cies, data on chromosomes and on a partial character- nant DNA emerging it was developed as an efficient ization of the genomes are already available [9,17–19]. system for heterologous gene expression instead, using The dimorphic yeasts A. adeninivorans and Y. lipolytica for the main part elements derived from methanol- represent more recently defined systems that have yet to metabolic pathway genes [25,26]. P. pastoris normally establish themselves but that demonstrate a promising exists in the vegetative haploid state
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