Functional identification of a delta8-sphingolipid desaturase from Borago officinalis

A - Papers appearing in refereed journals

Sperling, P., Libisch, B., Zahringer, U., Napier, J. A. and Heinz, E. 2001. Functional identification of a delta8-sphingolipid desaturase from Borago officinalis. Archives of Biochemistry and Biophysics. 388 (2), pp. 293-298.

AuthorsSperling, P., Libisch, B., Zahringer, U., Napier, J. A. and Heinz, E.
Abstract

The similarities between Δ12- and Δ15-fatty acyl desaturase sequences were used to construct degenerate primers for PCR experiments with cDNA transcribed from mRNA of developing borage seeds. Screening of a borage seed cDNA library with an amplified DNA fragment resulted in the isolation of a full-length cDNA corresponding to a deduced open-reading frame of 446 amino acids. The protein showed high similarity to plant Δ8-sphingolipid desaturases as well as to the Δ6-fatty acyl desaturase from Borago officinalis. The sequence is characterized by the presence of a N-terminal cytochrome b5 domain. Expression of this open-reading frame in Saccharomyces cerevisiae resulted in the formation of Δ8-trans/cis-phytosphingenines not present in wild-type cells, as shown by HPLC analysis of sphingoid bases as their dinitrophenyl derivatives. GLC-MS analysis of the methylated di-0-trimethylsilyl ether derivatives confirmed the presence of Δ8-stereoisomers of C18- and C20-phytosphingenine. Furthermore, Northern blotting showed that the gene encoding a stereo-unselective Δ8-sphingolipid desaturase is primarily expressed in young borage leaves.

Keywordscytochrome b5; desaturase; Borago officinalis; GLC-MS; phytosphingenine; reversed-phase HPLC; Saccharomyces cerevisiae; sphingolipids
Year of Publication2001
JournalArchives of Biochemistry and Biophysics
Journal citation388 (2), pp. 293-298
Digital Object Identifier (DOI)doi:10.1006/abbi.2001.2308
Open accessPublished as non-open access
Funder project or code415
503
Project: 3258
Output statusPublished
Publication dates
Online25 May 2002
Copyright licensePublisher copyright
ISSN0003-9861
PublisherElsevier Science Inc

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