Limits...
Novel lysophospholipid acyltransferase PLAT1 of Aurantiochytrium limacinum F26-b responsible for generation of palmitate-docosahexaenoate-phosphatidylcholine and phosphatidylethanolamine.

Abe E, Ikeda K, Nutahara E, Hayashi M, Yamashita A, Taguchi R, Doi K, Honda D, Okino N, Ito M - PLoS ONE (2014)

Bottom Line: The major source of DHA is fish oils but a recent increase in the global demand of DHA and decrease in fish stocks require a substitute.PLAT1 shows wide specificity for donor substrates as well as acceptor substrates in vitro, i.e, the enzyme can adopt lysophosphatidylcholine, lysophosphatidylethanolamine, lysophosphatidylserine and lysophosphatidylinositol as acceptor substrates, and 15:0/16:0-CoA and DHA-CoA as donor substrates.These results indicate that PLAT1 is the enzyme responsible for the generation of 16:0-DHA-PC and 16:0-DHA-PE in the thraustochytrid.

View Article: PubMed Central - PubMed

Affiliation: Department of Bioscience and Biotechnology, Graduate School of Bioresource and Bioenvironmental Sciences, Kyushu University, Fukuoka, Japan.

ABSTRACT
N-3 polyunsaturated fatty acids (PUFA), such as docosahexaenoic acid (DHA, 22:6n-3), have been reported to play roles in preventing cardiovascular diseases. The major source of DHA is fish oils but a recent increase in the global demand of DHA and decrease in fish stocks require a substitute. Thraustochytrids, unicellular marine protists belonging to the Chromista kingdom, can synthesize large amounts of DHA, and, thus, are expected to be an alternative to fish oils. DHA is found in the acyl chain(s) of phospholipids as well as triacylglycerols in thraustochytrids; however, how thraustochytrids incorporate DHA into phospholipids remains unknown. We report here a novel lysophospholipid acyltransferase (PLAT1), which is responsible for the generation of DHA-containing phosphatidylcholine and phosphatidylethanolamine in thraustochytrids. The PLAT1 gene, which was isolated from the genomic DNA of Aurantiochytrium limacinum F26-b, was expressed in Saccharomyces cerevisiae, and the FLAG-tagged recombinant enzyme was characterized after purification with anti-FLAG affinity gel. PLAT1 shows wide specificity for donor substrates as well as acceptor substrates in vitro, i.e, the enzyme can adopt lysophosphatidylcholine, lysophosphatidylethanolamine, lysophosphatidylserine and lysophosphatidylinositol as acceptor substrates, and 15:0/16:0-CoA and DHA-CoA as donor substrates. In contrast to the in vitro experiment, only lysophosphatidylcholine acyltransferase and lysophosphatidylethanolamine acyltransferase activities were decreased in plat1-knockout mutants, resulting in a decrease of 16:0-DHA-phosphatidylcholine (PC) [PC(38:6)] and 16:0-DHA-phosphatidylethanolamine (PE) [PE(38:6)], which are two major DHA-containing phospholipids in A. limacinum F26-b. However, the amounts of other phospholipid species including DHA-DHA-PC [PC(44:12)] and DHA-DHA-PE [PE(44:12)] were almost the same in plat-knockout mutants and the wild-type. These results indicate that PLAT1 is the enzyme responsible for the generation of 16:0-DHA-PC and 16:0-DHA-PE in the thraustochytrid.

Show MeSH

Related in: MedlinePlus

Alignment of PLAT1, mLPCAT1, and mLPCAT2.PLAT1 (this work), mLPCAT1 (LPCAT1 from mouse), and mLPCAT2 (LPCAT2 from mouse) sequences were aligned using GENETYX ver.8.2.2. The conserved amino acids are shown by white characters on a black background. The four conserved AGPAT motifs are indicated by red boxes. Two transmembrane regions, predicted by TMHMM server v. 2.0 (www.cbs.dtu.dk/services/TMHMM/), are underlined in blue. Three EF hand Ca2+-binding motifs, predicted by PROSITE (www.expasy.ch/prosite/), are indicated by green dashed-lines. ER-retaining motifs are indicated by red characters.
© Copyright Policy
Related In: Results  -  Collection

License
getmorefigures.php?uid=PMC4121067&req=5

pone-0102377-g002: Alignment of PLAT1, mLPCAT1, and mLPCAT2.PLAT1 (this work), mLPCAT1 (LPCAT1 from mouse), and mLPCAT2 (LPCAT2 from mouse) sequences were aligned using GENETYX ver.8.2.2. The conserved amino acids are shown by white characters on a black background. The four conserved AGPAT motifs are indicated by red boxes. Two transmembrane regions, predicted by TMHMM server v. 2.0 (www.cbs.dtu.dk/services/TMHMM/), are underlined in blue. Three EF hand Ca2+-binding motifs, predicted by PROSITE (www.expasy.ch/prosite/), are indicated by green dashed-lines. ER-retaining motifs are indicated by red characters.

Mentions: To identify the genes encoding LPLATs responsible for generating DHA-containing PLs in thraustochytrids, we searched the A. limacinum ATCC MYA-1381 genome database for sequence homology with previously reported LPLATs (Figure 2). We found seven candidate sequences in the database and designated them as PLAT1∼7, all of which belong to the AGPAT (LPAT) family. In this study, we cloned the PLAT1 gene (plat1) from the genomic DNA of A. limacinum F26-b, which is genealogically related to A. limacinum ATCC MYA-1381 (Figure 1).


Novel lysophospholipid acyltransferase PLAT1 of Aurantiochytrium limacinum F26-b responsible for generation of palmitate-docosahexaenoate-phosphatidylcholine and phosphatidylethanolamine.

Abe E, Ikeda K, Nutahara E, Hayashi M, Yamashita A, Taguchi R, Doi K, Honda D, Okino N, Ito M - PLoS ONE (2014)

Alignment of PLAT1, mLPCAT1, and mLPCAT2.PLAT1 (this work), mLPCAT1 (LPCAT1 from mouse), and mLPCAT2 (LPCAT2 from mouse) sequences were aligned using GENETYX ver.8.2.2. The conserved amino acids are shown by white characters on a black background. The four conserved AGPAT motifs are indicated by red boxes. Two transmembrane regions, predicted by TMHMM server v. 2.0 (www.cbs.dtu.dk/services/TMHMM/), are underlined in blue. Three EF hand Ca2+-binding motifs, predicted by PROSITE (www.expasy.ch/prosite/), are indicated by green dashed-lines. ER-retaining motifs are indicated by red characters.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4121067&req=5

pone-0102377-g002: Alignment of PLAT1, mLPCAT1, and mLPCAT2.PLAT1 (this work), mLPCAT1 (LPCAT1 from mouse), and mLPCAT2 (LPCAT2 from mouse) sequences were aligned using GENETYX ver.8.2.2. The conserved amino acids are shown by white characters on a black background. The four conserved AGPAT motifs are indicated by red boxes. Two transmembrane regions, predicted by TMHMM server v. 2.0 (www.cbs.dtu.dk/services/TMHMM/), are underlined in blue. Three EF hand Ca2+-binding motifs, predicted by PROSITE (www.expasy.ch/prosite/), are indicated by green dashed-lines. ER-retaining motifs are indicated by red characters.
Mentions: To identify the genes encoding LPLATs responsible for generating DHA-containing PLs in thraustochytrids, we searched the A. limacinum ATCC MYA-1381 genome database for sequence homology with previously reported LPLATs (Figure 2). We found seven candidate sequences in the database and designated them as PLAT1∼7, all of which belong to the AGPAT (LPAT) family. In this study, we cloned the PLAT1 gene (plat1) from the genomic DNA of A. limacinum F26-b, which is genealogically related to A. limacinum ATCC MYA-1381 (Figure 1).

Bottom Line: The major source of DHA is fish oils but a recent increase in the global demand of DHA and decrease in fish stocks require a substitute.PLAT1 shows wide specificity for donor substrates as well as acceptor substrates in vitro, i.e, the enzyme can adopt lysophosphatidylcholine, lysophosphatidylethanolamine, lysophosphatidylserine and lysophosphatidylinositol as acceptor substrates, and 15:0/16:0-CoA and DHA-CoA as donor substrates.These results indicate that PLAT1 is the enzyme responsible for the generation of 16:0-DHA-PC and 16:0-DHA-PE in the thraustochytrid.

View Article: PubMed Central - PubMed

Affiliation: Department of Bioscience and Biotechnology, Graduate School of Bioresource and Bioenvironmental Sciences, Kyushu University, Fukuoka, Japan.

ABSTRACT
N-3 polyunsaturated fatty acids (PUFA), such as docosahexaenoic acid (DHA, 22:6n-3), have been reported to play roles in preventing cardiovascular diseases. The major source of DHA is fish oils but a recent increase in the global demand of DHA and decrease in fish stocks require a substitute. Thraustochytrids, unicellular marine protists belonging to the Chromista kingdom, can synthesize large amounts of DHA, and, thus, are expected to be an alternative to fish oils. DHA is found in the acyl chain(s) of phospholipids as well as triacylglycerols in thraustochytrids; however, how thraustochytrids incorporate DHA into phospholipids remains unknown. We report here a novel lysophospholipid acyltransferase (PLAT1), which is responsible for the generation of DHA-containing phosphatidylcholine and phosphatidylethanolamine in thraustochytrids. The PLAT1 gene, which was isolated from the genomic DNA of Aurantiochytrium limacinum F26-b, was expressed in Saccharomyces cerevisiae, and the FLAG-tagged recombinant enzyme was characterized after purification with anti-FLAG affinity gel. PLAT1 shows wide specificity for donor substrates as well as acceptor substrates in vitro, i.e, the enzyme can adopt lysophosphatidylcholine, lysophosphatidylethanolamine, lysophosphatidylserine and lysophosphatidylinositol as acceptor substrates, and 15:0/16:0-CoA and DHA-CoA as donor substrates. In contrast to the in vitro experiment, only lysophosphatidylcholine acyltransferase and lysophosphatidylethanolamine acyltransferase activities were decreased in plat1-knockout mutants, resulting in a decrease of 16:0-DHA-phosphatidylcholine (PC) [PC(38:6)] and 16:0-DHA-phosphatidylethanolamine (PE) [PE(38:6)], which are two major DHA-containing phospholipids in A. limacinum F26-b. However, the amounts of other phospholipid species including DHA-DHA-PC [PC(44:12)] and DHA-DHA-PE [PE(44:12)] were almost the same in plat-knockout mutants and the wild-type. These results indicate that PLAT1 is the enzyme responsible for the generation of 16:0-DHA-PC and 16:0-DHA-PE in the thraustochytrid.

Show MeSH
Related in: MedlinePlus