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The Oxidosqualene Cyclase from the Oomycete Saprolegnia parasitica Synthesizes Lanosterol as a Single Product
Stockholm University, Faculty of Science, Department of Ecology, Environment and Plant Sciences. KTH Royal Institute of Technology, Sweden.ORCID iD: 0000-0002-9945-8059
2016 (English)In: Frontiers in Microbiology, ISSN 1664-302X, E-ISSN 1664-302X, Vol. 7, 1802Article in journal (Refereed) Published
Abstract [en]

The first committed step of sterol biosynthesis is the cyclisation of 2,3-oxidosqualene to form either lanosterol (LA) or cycloartenol (CA). This is catalyzed by an oxidosqualene cyclase (OSC). LA and CA are subsequently converted into various sterols by a series of enzyme reactions. The specificity of the OSC therefore determines the final composition of the end sterols of an organism. Despite the functional importance of OSCs, the determinants of their specificity are not well understood. In sterol-synthesizing oomycetes, recent bioinformatics, and metabolite analysis suggest that LA is produced. However, this catalytic activity has never been experimentally demonstrated. Here, we show that the OSC of the oomycete Saprolegnia parasitica, a severe pathogen of salmonid fish, has an uncommon sequence in a conserved motif important for specificity. We present phylogenetic analysis revealing that this sequence is common to sterol-synthesizing oomycetes, as well as some plants, and hypothesize as to the evolutionary origin of some microbial sequences. We also demonstrate for the first time that a recombinant form of the OSC from S. parasitica produces LA exclusively. Our data pave the way for a detailed structural characterization of the protein and the possible development of specific inhibitors of oomycete OSCs for disease control in aquaculture.

Place, publisher, year, edition, pages
2016. Vol. 7, 1802
Keyword [en]
lanosterol biosynthesis, oomycete, Saprolegnia parasitica, oxidosqualene cyclase, sterols
National Category
Biological Sciences
Research subject
Plant Physiology
Identifiers
URN: urn:nbn:se:su:diva-136547DOI: 10.3389/fmicb.2016.01802ISI: 000388698600001OAI: oai:DiVA.org:su-136547DiVA: diva2:1055146
Projects
No
Available from: 2016-12-11 Created: 2016-12-11 Last updated: 2017-01-09Bibliographically approved
In thesis
1. Analysis of sterol metabolism in the pathogenic oomycetes Saprolegnia parasitica and Phytophthora infestans
Open this publication in new window or tab >>Analysis of sterol metabolism in the pathogenic oomycetes Saprolegnia parasitica and Phytophthora infestans
2016 (English)Doctoral thesis, comprehensive summary (Other academic)
Abstract [en]

The primary objective of this thesis was to investigate the sterol metabolism of two pathogenic oomycetes, specifically the processes of sterol synthesis and sterol acquisition in the fish pathogen Saprolegnia parasitica (Saprolegniales) and the plant pathogen Phytophthora infestans (Peronosporales). Furthermore, the effects of steroidal glycoalkaloids from Solanaceous plants, on P. infestans, were examined. The improved understanding of these processes should help to identify approaches for the identification of new oomycete inhibitors targeting sterol metabolism in agriculture and aquaculture farming systems, and to guide plant-breeding strategies to defend solanaceous plants against oomycetes.

For these reasons, the molecular basis of the metabolic pathways of sterol synthesis and/or sterol acquisition was investigated. Sterols are derived from isoprenoids and indispensable in various biological processes. Our biochemical investigation of an oxidosqualene cyclase revealed that sterol synthesis in S. parasitica begins with the formation of lanosterol (Paper I), and a reconstruction of the complete sterol synthesis pathway to the final compound, fucosterol, in S. parasitica was performed using bioinformatics (Paper II). Complementary to this work, the extent to which P. infestans, which is incapable of de novo sterol synthesis, is able to modify exogenously provided sterols was investigated by determining the growth impact of various sterol supplements in the growth media (Paper II). 

Building on the sterol investigations, the solanaceous sterol derivatives from the glycoalkaloid family were analysed. These compounds contain both a steroidal and a carbohydrate (glycan) moiety. Data obtained by feeding various deuterium-labeled sterols to potato shoots, supported the theory that steroidal glycoalkaloids in Solanum tuberosum are produced from cholesterol (Paper III).  Since these steroidal glycoalkaloids are thought to play a role in plant defense, their physiological effects on P. infestans were investigated (Paper IV). Unexpectedly we found that non-glycosylated steroidal alkaloids had a greater inhibitory effect than steroidal glycoalkaloids.  Steroidal glycoalkaloids derived from other Solanaceous species exhibited different physiological effects on the growth of P. infestans

This research was conducted on two oomycete species belonging to the Saprolegniales and Peronosporales orders, hence the results presented are likely to be representative of each of these two oomycete orders.

Place, publisher, year, edition, pages
Stockholm: Department of Ecology, Environment and Plant Sciences, Stockholm University, 2016. 59 p.
National Category
Biochemistry and Molecular Biology
Research subject
Plant Physiology
Identifiers
urn:nbn:se:su:diva-136551 (URN)978-91-7649-652-7 (ISBN)978-91-7649-653-4 (ISBN)
Public defence
2017-01-27, sal P216, Arrheniuslaboratorierna, Svante Arrhenius väg 20, Stockholm, 10:00 (English)
Opponent
Supervisors
Note

At the time of the doctoral defense, the following papers were unpublished and had a status as follows: Paper 2: Manuscript. Paper 4: Manuscript.

Available from: 2017-01-02 Created: 2016-12-11 Last updated: 2017-01-27Bibliographically approved

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