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J. Biol. Chem., Vol. 281, Issue 31, 21988-21997, August 4, 2006

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Identification and Functional Characterization of the Moss Physcomitrella patens ⁵-Desaturase Gene Involved in Arachidonic and Eicosapentaenoic Acid Biosynthesis^*

Songsri Kaewsuwan, Edgar B. Cahoon, Pierre-François Perroud^¶, Chanpen Wiwat^||, Nathinee Panvisavas^**, Ralph S. Quatrano, Supported by Washington University in St. Louis^¶1, David J. Cove^¶2, and Nuntavan Bunyapraphatsara³

From the Departments of Pharmacognosy and ^||Microbiology, Faculty of Pharmacy, Mahidol University, 447 Sri-Ayudhya Road, Ratchathewi, Bangkok 10400, Thailand, ^**Department of Plant Sciences, Faculty of Sciences, Mahidol University, Rama VI Road, Bangkok 10400, Thailand, United States Department of Agriculture Agricultural Research Service, Plant Genetics Research Unit, Donald Danforth Plant Science Center, St. Louis, Missouri 63132, and the ^¶Department of Biology, Washington University in St. Louis, S Louis, Missouri 63130-4899

The moss Physcomitrella patens contains high levels of arachidonic acid and lesser amounts of eicosapentaenoic acid. Here we report the identification and characterization of a ⁵-desaturase from P. patens that is associated with the synthesis of these fatty acids. A full-length cDNA for this desaturase was identified by data base searches based on homology to sequences of known ⁵-desaturase cDNAs from fungal and algal species. The resulting P. patens cDNA encodes a 480-amino acid polypeptide that contains a predicted N-terminal cytochrome b₅-like domain as well as three histidine-rich domains. Expression of the enzyme in Saccharomyces cerevisiae resulted in the production of the ⁵-containing fatty acid arachidonic acid in cells that were provided di-homo--linolenic acid. In addition, the expressed enzyme generated ⁵-desaturation products with the C20 substrates -6 eicosadienoic and -3 eicosatrienoic acids, but no products were detected with the C18 fatty acid linoleic and -linolenic acids or with the C22 fatty acid adrenic and docosapentaenoic acids. When the corresponding P. patens genomic sequence was disrupted by replacement through homologous recombination, a dramatic alteration in the fatty acid composition was observed, i.e. an increase in di-homo--linolenic and eicosatetraenoic acids accompanied by a concomitant disappearance of the ⁵-fatty acid arachidonic and eicosapentaenoic acids. In addition, overexpression of the P. patens cDNA in protoplasts isolated from a disrupted line resulted in the restoration of arachidonic acid synthesis.

Received for publication, March 30, 2006 , and in revised form, May 12, 2006.

The amino acid sequence of this protein can be accessed through NCBI Protein Database under NCBI accession number CAI58861 [GenBank] .

^* This work was based on a doctoral study by S. K. in the Faculty of Pharmacy, Mahidol University supported by the Royal Golden Jubilee Ph.D. research scholarship and the Current Program Sponsor funds of Washington University in St. Louis. The costs of publication of this article were defrayed in part by the payment of page charges. This article must therefore be hereby marked "advertisement" in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.

² Supported by the Leverhulme Trust and George W. and Irene K. Freiberg funds of Washington University in St. Louis.

¹ To whom correspondence may be addressed: Dept. of Biology, Washington University in St. Louis, One Brookings Dr., St. Louis, MO 63130-4899. Tel.: 314-935-6850; E-mail: rsq{at}biology.wustl.edu. ³ To whom correspondence may be addressed. Tel.: 66-2-644-4566; Fax: 66-2-644-4566; E-mail: pynby{at}mahidol.ac.th.

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This article has been cited by other articles:

				D. J. Cove, P.-F. Perroud, A. J. Charron, S. F. McDaniel, A. Khandelwal, and R. S. Quatrano The Moss Physcomitrella patens: A Novel Model System for Plant Development and Genomic Studies CSH Protocols, February 1, 2009; 2009(2): pdb.emo115 - pdb.emo115. [Abstract] [Full Text]