Isolation and characterization of cerebrosides of the hydrocarbon‐assimilating yeast Yarrowia lipolytica

Abstract

Yarrowia lipolytica yeast was grown batchwise on n-hexadecane as the carbon and energy source. Two cerebroside species were quantitatively isolated from sphingolipid fractions of total lipids by a combination of column chromatography and preparative high-performance thin-layer chromatography. The cerebroside content accounted for 1.3% of the total cell lipids. Glucose was detected as the sole sugar constituent in cerebrosides. The fatty acid composition of cerebrosides was characterized by a high proportion of hydroxylated long-chain saturated fatty acids. The major fatty acids were h16:0 and 16:0. The long-chain bases composition shows a preponderance of trihydroxy bases and a small amount of dihydroxy bases. The striking finding was a high proportion of 19-phytosphingosine.

Significance of theKlLAC1gene in glucosylceramide production byKluyveromyces lactis

“…LAG1p, the longevity-assurance gene 1 (D'Mello et al, 1994), or LAC1p, the longevity-assurance gene cognate 1 (Jiang et al, 1998), associated with LIP1p, the LAG1p/LAC1p interaction protein (Kageyama-Yahara & Riezman, 2006). Not all of the yeast species can synthesize glucosylceramide (Saito et al, 2006), and those accumulating glucosylceramide (Matsubara et al, 1987;Takakuwa et al, 2002Takakuwa et al, , 2005aRupčić et al, 2004) were expected to possess LAC1p, similar to KlLAC1p. However, KlLAC1p from K. lactis has shown to be the enzyme that strictly produces ceramide composed of the sphingoid base and C 18 fatty acid as the precursor of glucosylceramide, as indicated by the observations of the overexpression and disruption of the KlLAC1 gene in K. lactis and the heterologous expression of the KlLAC1 gene in S. cerevisiae.…”

“…However, KlLAC1p from K. lactis has shown to be the enzyme that strictly produces ceramide composed of the sphingoid base and C 18 fatty acid as the precursor of glucosylceramide, as indicated by the observations of the overexpression and disruption of the KlLAC1 gene in K. lactis and the heterologous expression of the KlLAC1 gene in S. cerevisiae. Not all of the yeast species can synthesize glucosylceramide (Saito et al, 2006), and those accumulating glucosylceramide (Matsubara et al, 1987;Takakuwa et al, 2002Takakuwa et al, , 2005aRupčić et al, 2004) were expected to possess LAC1p, similar to KlLAC1p. When the amino acid sequences of (Dietrich et al, 2004), Candida albicans LAC1p (XP_716595) and LAG1p (XP_717940) (Jones et al, 2004), Candida glabrata LAC1p (XP_449791) and LAG1p (XP_448344), Debaryomyces hansenii LAC1p (XP_456760) and LAG1p (XP_460173), Kluyveromyces lactis LAC1p (XP_454318) and LAG1p (XP_452132) (Dujon et al, 2004), Saccharomyces cerevisiae LAC1p (NP_012917) and LAG1p (NP_011860) (Goffeau et al, 1996), Vanderwaltozyma polyspora LAC1p (XP_001647376) and LAG1p (XP_001644693) (Scannell et al, 2007), Yarrowia lipolytica LAC1p (XP_502386) and LAG1p (XP_505079) (Dujon et al, 2004), and human LASS1p (NP_067090) (Jiang et al, 1998), LASS2p (NP_071358) (Pan et al, 2001), LASS3p (AAH34970), LASS4p (AAH09828), LASS5p (AAH32565) (Strausberg et al, 2002) and LASS6p (NP_982288) (Harrington et al, 2001).…”