Identification and expression of mammalian long-chain PUFA elongation enzymes.
In mammalian cells, Sprecher has proposed that the synthesis of long-chain PUFA from the 20-carbon substrates involves two consecutive elongation steps, a delta6-desaturation step followed by retroconversion (Sprecher, H., Biochim. Biophys. Acta 1486, 219-231, 2000). We searched the database using the translated sequence of human elongase ELOVL5, whose encoded enzyme elongates monounsaturated and polyunsaturated FA, as a query to identify the enzyme(s) involved in elongation of very long chain PUFA. The database search led to the isolation of two cDNA clones from human and mouse. These clones displayed deduced amino acid sequences that had 56.4 and 58% identity, respectively, to that of ELOVL5. The open reading frame of the human clone (ELOVL2) encodes a 296-amino acid peptide, whereas the mouse clone (Elovl2) encodes a 292-amino acid peptide. Expression of these open reading frames in baker's yeast, Saccharomyces cerevisiae, demonstrated that the encoded proteins were involved in the elongation of both 20- and 22-carbon long-chain PUFA, as determined by the conversion of 20:4n-6 to 22:4n-6, 22:4n-6 to 24:4n-6, 20:5n-3 to 22:5n-3, and 22:5n-3 to 24:5n-3. The elongation activity of the mouse Elovl2 was further demonstrated in the transformed mouse L cells incubated with long-chain (C20- and C22-carbon) n-6 and n-3 PUFA substrates by the significant increase in the levels of 24:4n-6 and 24:5n-3, respectively. This report demonstrates the isolation and identification of two mammalian genes that encode very long chain PUFA specific elongation enzymes in the Sprecher pathway for DHA synthesis.
Leonard
Amanda E
AE
Ross Products Division, Abbott Laboratories, Columbus, Ohio 43215, USA.
Kelder
Bruce
B
Bobik
Emil G
EG
Chuang
Lu-Te
LT
Lewis
Christopher J
CJ
Kopchick
John J
JJ
Mukerji
Pradip
P
Huang
Yung-Sheng
YS
D000123:Acetyltransferases / Q000235:genetics* / Q000378:metabolism*; D000595:Amino Acid Sequence; D000818:Animals; D015152:Blotting, Northern; D003001:Cloning, Molecular; D018076:DNA, Complementary / Q000235:genetics; D044943:Fatty Acid Desaturases / Q000378:metabolism; D000081031:Fatty Acid Elongases; D015525:Fatty Acids, Omega-3 / Q000096:biosynthesis*; D043371:Fatty Acids, Omega-6; D005231:Fatty Acids, Unsaturated / Q000096:biosynthesis*; D008401:Gas Chromatography-Mass Spectrometry; D005822:Genetic Vectors / Q000235:genetics; D006801:Humans; D051379:Mice; D008969:Molecular Sequence Data; D016366:Open Reading Frames / Q000235:genetics; D009928:Organ Specificity; D012441:Saccharomyces cerevisiae / Q000235:genetics / Q000378:metabolism; D017386:Sequence Homology, Amino Acid
D018076:DNA, Complementary; C000630967:ELOVL2 protein, human; C000630951:ELOVL2 protein, mouse; C000630954:ELOVL5 protein, human; D015525:Fatty Acids, Omega-3; D043371:Fatty Acids, Omega-6; D005231:Fatty Acids, Unsaturated; D044943:Fatty Acid Desaturases; D000123:Acetyltransferases; D000081031:Fatty Acid Elongases