The Experts below are selected from a list of 264 Experts worldwide ranked by ideXlab platform
Michael Schlame - One of the best experts on this subject based on the ideXlab platform.
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the basis for acyl specificity in the tafazzin reaction
Journal of Biological Chemistry, 2017Co-Authors: Michael Schlame, Yang XuAbstract:: Tafazzin is a mitochondrial enzyme that transfers fatty acids from phospholipids to lysophospholipids. Mutations in tafazzin cause abnormal molecular species of cardiolipin and the clinical phenotype of Barth syndrome. However, the mechanism by which tafazzin creates acyl specificity has been controversial. We have shown that the lipid phase state can produce acyl specificity in the tafazzin reaction, but others have reported that tafazzin itself carries enzymatic specificity. To resolve this issue, we replicated and expanded the controversial experiments, i.e. the transfer of different acyl groups from phosphatidylcholine to monolysocardiolipin by yeast tafazzin. Our data show that this reaction requires the presence of detergent and does not take place in liposomes but in mixed micelles. To separate thermodynamic (lipid-dependent) from kinetic (enzyme-dependent) parameters, we followed the accumulation of cardiolipin during the reaction from the initial state to the equilibrium state. The Transacylation rates of different acyl groups varied over 2 orders of magnitude and correlated tightly with the concentration of cardiolipin in the equilibrium state (lipid-dependent parameter). In contrast, the rates by which different Transacylations approached the equilibrium state were very similar (enzyme-dependent parameter). Furthermore, we found that tafazzin catalyzes the remodeling of cardiolipin by combinations of forward and reverse Transacylations, essentially creating an equilibrium distribution of acyl groups. These data strongly support the idea that the acyl specificity of the tafazzin reaction results from the physical properties of lipids.
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the physical state of lipid substrates provides Transacylation specificity for tafazzin
Nature Chemical Biology, 2012Co-Authors: Michael Schlame, Bob Berno, Devrim Acehan, David L Stokes, Salvatore Valvo, Mindong Ren, Richard M EpandAbstract:Cardiolipin is a mitochondrial phospholipid with a characteristic acyl chain composition that depends on the function of tafazzin, a phospholipid-lysophospholipid transacylase, although the enzyme itself lacks acyl specificity. We incubated isolated tafazzin with various mixtures of phospholipids and lysophospholipids, characterized the lipid phase by (31)P-NMR and measured newly formed molecular species by MS. Substantial Transacylation was observed only in nonbilayer lipid aggregates, and the substrate specificity was highly sensitive to the lipid phase. In particular, tetralinoleoyl-cardiolipin, a prototype molecular species, formed only under conditions that favor the inverted hexagonal phase. In isolated mitochondria, <1% of lipids participated in Transacylations, suggesting that the action of tafazzin was limited to privileged lipid domains. We propose that tafazzin reacts with non-bilayer-type lipid domains that occur in curved or hemifused membrane zones and that acyl specificity is driven by the packing properties of these domains.
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reconstitution of acyl specific phospholipid remodeling by purified tafazzin in vitro
Biophysical Journal, 2012Co-Authors: Michael Schlame, Bob Berno, Devrim Acehan, Yang Xu, David L Stokes, Richard M EpandAbstract:Cardiolipin is a mitochondrial phospholipid with a unique composition and distribution of acyl groups. The cardiolipin composition depends on tafazzin, a phospholipid-lysophospholipid transacylase, although the enzyme itself lacks acyl specificity. We incubated isolated tafazzin with various mixtures of phospholipids and lysophospholipids, characterized the lipid phase state by 31P-NMR, and measured newly formed molecular species by mass spectrometry. Significant Transacylation activity was observed only in non-bilayer lipid aggregates, in which lipids had a low packing order. The lipid phase state profoundly affected the substrate specificity of the tafazzin reaction. In particular, tetralinoleoyl-cardiolipin, a prototype molecular species, formed only under conditions that favor the inverted hexagonal phase. In isolated mitochondria, less than 2 percent of lipids participated in Transacylations, suggesting that tafazzin acts only on privileged lipid domains. We propose that tafazzin reacts with non-bilayer lipids in mitochondria and that acyl specificity arises from spontaneous self-organization of these domains.View Large Image | View Hi-Res Image | Download PowerPoint Slide
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Formation of molecular species of mitochondrial cardiolipin 2. A mathematical model of pattern formation by phospholipid Transacylation
Biochimica et Biophysica Acta, 2009Co-Authors: Michael SchlameAbstract:Formation of the unique molecular species of mitochondrial cardiolipin requires tafazzin, a transacylase that exchanges acyl groups between phospholipid molecular species without strict specificity for acyl groups, head groups, or carbon positions. However, it is not known whether phospholipid Transacylations can cause the accumulation of specific fatty acids in cardiolipin. Here, a model is shown in linear algebra representation, in which acyl specificity emerges from the Transacylation equilibrium of multiple molecular species, provided that different species have different free energies. The model defines the conditions and energy terms, under which Transacylations may generate the characteristic composition of mitochondrial cardiolipin. It is concluded that acyl-specific cardiolipin patterns could arise from phospholipid Transacylations in the tafazzin domain, even if tafazzin itself does not have substrate specificity.
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the enzymatic function of tafazzin
Journal of Biological Chemistry, 2006Co-Authors: Yang Xu, Ashim Malhotra, Michael SchlameAbstract:Abstract Tafazzin is a putative enzyme that is involved in cardiolipin metabolism, it may carry mutations responsible for Barth syndrome. To identify the biochemical reaction catalyzed by tafazzin, we expressed the full-length isoform of Drosophila melanogaster tafazzin in a baculovirus-Sf9 insect cell system. Tafazzin expression induced a new enzymatic function in Sf9 cell mitochondria, namely 1-palmitoyl-2-[14C]linoleoyl-phosphatidylcholine:monolysocardiolipin linoleoyltransferase. We also found evidence for the reverse reaction, because tafazzin expression caused transfer of acyl groups from phospholipids to 1-[14C]palmitoyl-2-lyso-phosphatidylcholine. An affinity-purified tafazzin construct, tagged with the maltose-binding protein, catalyzed both forward and reverse Transacylations between cardiolipin and phosphatidylcholine, but was unable to utilize CoA or acyl-CoA as substrates. Whereas tafazzin supported Transacylations between various phospholipid-lysophospholipid pairs, it showed the highest rate for the phosphatidylcholine-cardiolipin Transacylation. Transacylation activities were about 10-fold higher for linoleoyl groups than for oleoyl groups, and they were negligible for arachidonoyl groups. The data show that Drosophila tafazzin is a CoA-independent, acyl-specific phospholipid transacylase with substrate preference for cardiolipin and phosphatidylcholine.
Takayuki Sugiura - One of the best experts on this subject based on the ideXlab platform.
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Coenzyme-A-Independent Transacylation System; Possible Involvement of Phospholipase A2 in Transacylation.
Biology, 2017Co-Authors: Atsushi Yamashita, Yasuhiro Hayashi, Naoki Matsumoto, Yoko Nemoto-sasaki, Takanori Koizumi, Yusuke Inagaki, Saori Oka, Takashi Tanikawa, Takayuki SugiuraAbstract:The coenzyme A (CoA)-independent Transacylation system catalyzes fatty acid transfer from phospholipids to lysophospholipids in the absence of cofactors such as CoA. It prefers to use C20 and C22 polyunsaturated fatty acids such as arachidonic acid, which are esterified in the glycerophospholipid at the sn-2 position. This system can also acylate alkyl ether-linked lysophospholipids, is involved in the enrichment of arachidonic acid in alkyl ether-linked glycerophospholipids, and is critical for the metabolism of eicosanoids and platelet-activating factor. Despite their importance, the enzymes responsible for these reactions have yet to be identified. In this review, we describe the features of the Ca2+-independent, membrane-bound CoA-independent Transacylation system and its selectivity for arachidonic acid. We also speculate on the involvement of phospholipase A2 in the CoA-independent Transacylation reaction.
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acyltransferases and transacylases that determine the fatty acid composition of glycerolipids and the metabolism of bioactive lipid mediators in mammalian cells and model organisms
Progress in Lipid Research, 2014Co-Authors: Atsushi Yamashita, Keizo Waku, Yasuhiro Hayashi, Saori Oka, Takashi Tanikawa, Yoko Nemotosasaki, Makoto Ito, Takayuki SugiuraAbstract:Abstract Over one hundred different phospholipid molecular species are known to be present in mammalian cells and tissues. Fatty acid remodeling systems for phospholipids including acyl-CoA:lysophospholipid acyltransferases, CoA-dependent and CoA-independent Transacylation systems, are involved in the biosynthesis of these molecular species. Acyl-CoA:lysophospholipid acyltransferase system is involved in the synthesis of phospholipid molecular species containing sn-1 saturated and sn-2 unsaturated fatty acids. The CoA-dependent Transacylation system catalyzes the transfer of fatty acids esterified in phospholipids to lysophospholipids in the presence of CoA without the generation of free fatty acids. The CoA-dependent Transacylation reaction in the rat liver exhibits strict fatty acid specificity, i.e., three types of fatty acids (20:4, 18:2 and 18:0) are transferred. On the other hand, CoA-independent transacylase catalyzes the transfer of C20 and C22 polyunsaturated fatty acids from diacyl phospholipids to various lysophospholipids, especially ether-containing lysophospholipids, in the absence of any cofactors. CoA-independent transacylase is assumed to be involved in the accumulation of PUFA in ether-containing phospholipids. These enzymes are involved in not only the remodeling of fatty acids, but also the synthesis and degradation of some bioactive lipids and their precursors. In this review, recent progresses in acyltransferase research including the identification of the enzyme’s genes are described.
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subcellular localization and lysophospholipase Transacylation activities of human group ivc phospholipase a2 cpla2γ
Biochimica et Biophysica Acta, 2009Co-Authors: Atsushi Yamashita, Ken Tanaka, Ryo Kamata, Tsukasa Kumazawa, Naotaka Suzuki, Hiroki Koga, Keizo Waku, Takayuki SugiuraAbstract:Abstract cPLA2γ was identified as an ortholog of cPLA2α, which is a key enzyme in eicosanoid production. cPLA2γ was reported to be located in endoplasmic reticulum (ER) and mitochondria and to have lysophospholipase activity beside phospholipase A2 (PLA2) activity. However, subcellular localization, mechanism of membrane binding, regulation and physiological function have not been fully established. In the present study, we examined the subcellular localization and enzymatic properties of cPLA2γ with C-terminal FLAG-tag. We found that cPLA2γ was located not only in ER but also mitochondria even in the absence of the prenylation. Purified recombinant cPLA2γ catalyzed an acyltransferase reaction from one molecule of lysophosphatidylcholine (LPC) to another, forming phosphatidylcholine (PC). LPC or lysophosphatidylethanolamine acted as acyl donor and acceptor, but lysophosphatidylserine, lysophosphatidylinositol and lysophosphatidic acid (LPA) did not. PC and phosphatidylethanolamine (PE) also acted as weak acyl donors. Reaction conditions changed the balance of lysophospholipase and Transacylation activities, with addition of LPA/PA, pH > 8, and elevated temperature markedly increasing Transacylation activity; this suggests that lysophospholipase/Transacylation activities of cPLA2γ may be regulated by various factors. As lysophospholipids are known to accumulate in ischemia heart and to induce arryhthmia, the cPLA2γ that is abundant in heart may have a protective role through clearance of lysophospholipids by its Transacylation activity.
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Subcellular localization and lysophospholipase/Transacylation activities of human group IVC phospholipase A2 (cPLA2γ)
Biochimica et biophysica acta, 2009Co-Authors: Atsushi Yamashita, Ken Tanaka, Ryo Kamata, Tsukasa Kumazawa, Naotaka Suzuki, Hiroki Koga, Keizo Waku, Takayuki SugiuraAbstract:Abstract cPLA2γ was identified as an ortholog of cPLA2α, which is a key enzyme in eicosanoid production. cPLA2γ was reported to be located in endoplasmic reticulum (ER) and mitochondria and to have lysophospholipase activity beside phospholipase A2 (PLA2) activity. However, subcellular localization, mechanism of membrane binding, regulation and physiological function have not been fully established. In the present study, we examined the subcellular localization and enzymatic properties of cPLA2γ with C-terminal FLAG-tag. We found that cPLA2γ was located not only in ER but also mitochondria even in the absence of the prenylation. Purified recombinant cPLA2γ catalyzed an acyltransferase reaction from one molecule of lysophosphatidylcholine (LPC) to another, forming phosphatidylcholine (PC). LPC or lysophosphatidylethanolamine acted as acyl donor and acceptor, but lysophosphatidylserine, lysophosphatidylinositol and lysophosphatidic acid (LPA) did not. PC and phosphatidylethanolamine (PE) also acted as weak acyl donors. Reaction conditions changed the balance of lysophospholipase and Transacylation activities, with addition of LPA/PA, pH > 8, and elevated temperature markedly increasing Transacylation activity; this suggests that lysophospholipase/Transacylation activities of cPLA2γ may be regulated by various factors. As lysophospholipids are known to accumulate in ischemia heart and to induce arryhthmia, the cPLA2γ that is abundant in heart may have a protective role through clearance of lysophospholipids by its Transacylation activity.
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Reverse reaction of lysophosphatidylinositol acyltransferase. Functional reconstitution of coenzyme A-dependent Transacylation system.
The Journal of biological chemistry, 2003Co-Authors: Atsushi Yamashita, Ryo Kamata, Masanobu Watanabe, Kazuaki Sato, Tomoyuki Miyashita, Tomonari Nagatsuka, Hironori Kondo, Norikazu Kawagishi, Hiroki Nakanishi, Takayuki SugiuraAbstract:CoA-dependent Transacylation activity in microsomes catalyzes the transfer of fatty acid between phospholipids and lysophospholipids in the presence of CoA without the generation of free fatty acid. We examined the mechanism of the Transacylation system using partially purified acyl-CoA:lysophosphatidylinositol (LPI) acyltransferase (LPIAT) from rat liver microsomes to test our hypothesis that both the reverse and forward reactions of acyl-CoA:lysophospholipid acyltransferases are involved in the CoA-dependent Transacylation process. The purified LPIAT fraction exhibited ATP-independent acyl-CoA synthetic activity and CoA-dependent LPI generation from PI, suggesting that LPIAT could operate in reverse to form acyl-CoA and LPI. CoA-dependent acylation of LPI by the purified LPIAT fraction required PI as the acyl donor. In addition, the combination of purified LPIAT and recombinant lysophosphatidic acid acyltransferase could reconstitute CoA-dependent Transacylation between PI and phosphatidic acid. These results suggest that the CoA-dependent Transacylation system consists of the following: 1) acyl-CoA synthesis from phospholipid through the reverse action of acyl-CoA:lysophospholipid acyltransferases; and 2) transfer of fatty acyl moiety from the newly formed acyl-CoA to lysophospholipid through the forward action of acyl-CoA:lysophospholipid acyltransferases.
Atsushi Yamashita - One of the best experts on this subject based on the ideXlab platform.
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Coenzyme-A-Independent Transacylation System; Possible Involvement of Phospholipase A2 in Transacylation.
Biology, 2017Co-Authors: Atsushi Yamashita, Yasuhiro Hayashi, Naoki Matsumoto, Yoko Nemoto-sasaki, Takanori Koizumi, Yusuke Inagaki, Saori Oka, Takashi Tanikawa, Takayuki SugiuraAbstract:The coenzyme A (CoA)-independent Transacylation system catalyzes fatty acid transfer from phospholipids to lysophospholipids in the absence of cofactors such as CoA. It prefers to use C20 and C22 polyunsaturated fatty acids such as arachidonic acid, which are esterified in the glycerophospholipid at the sn-2 position. This system can also acylate alkyl ether-linked lysophospholipids, is involved in the enrichment of arachidonic acid in alkyl ether-linked glycerophospholipids, and is critical for the metabolism of eicosanoids and platelet-activating factor. Despite their importance, the enzymes responsible for these reactions have yet to be identified. In this review, we describe the features of the Ca2+-independent, membrane-bound CoA-independent Transacylation system and its selectivity for arachidonic acid. We also speculate on the involvement of phospholipase A2 in the CoA-independent Transacylation reaction.
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acyltransferases and transacylases that determine the fatty acid composition of glycerolipids and the metabolism of bioactive lipid mediators in mammalian cells and model organisms
Progress in Lipid Research, 2014Co-Authors: Atsushi Yamashita, Keizo Waku, Yasuhiro Hayashi, Saori Oka, Takashi Tanikawa, Yoko Nemotosasaki, Makoto Ito, Takayuki SugiuraAbstract:Abstract Over one hundred different phospholipid molecular species are known to be present in mammalian cells and tissues. Fatty acid remodeling systems for phospholipids including acyl-CoA:lysophospholipid acyltransferases, CoA-dependent and CoA-independent Transacylation systems, are involved in the biosynthesis of these molecular species. Acyl-CoA:lysophospholipid acyltransferase system is involved in the synthesis of phospholipid molecular species containing sn-1 saturated and sn-2 unsaturated fatty acids. The CoA-dependent Transacylation system catalyzes the transfer of fatty acids esterified in phospholipids to lysophospholipids in the presence of CoA without the generation of free fatty acids. The CoA-dependent Transacylation reaction in the rat liver exhibits strict fatty acid specificity, i.e., three types of fatty acids (20:4, 18:2 and 18:0) are transferred. On the other hand, CoA-independent transacylase catalyzes the transfer of C20 and C22 polyunsaturated fatty acids from diacyl phospholipids to various lysophospholipids, especially ether-containing lysophospholipids, in the absence of any cofactors. CoA-independent transacylase is assumed to be involved in the accumulation of PUFA in ether-containing phospholipids. These enzymes are involved in not only the remodeling of fatty acids, but also the synthesis and degradation of some bioactive lipids and their precursors. In this review, recent progresses in acyltransferase research including the identification of the enzyme’s genes are described.
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subcellular localization and lysophospholipase Transacylation activities of human group ivc phospholipase a2 cpla2γ
Biochimica et Biophysica Acta, 2009Co-Authors: Atsushi Yamashita, Ken Tanaka, Ryo Kamata, Tsukasa Kumazawa, Naotaka Suzuki, Hiroki Koga, Keizo Waku, Takayuki SugiuraAbstract:Abstract cPLA2γ was identified as an ortholog of cPLA2α, which is a key enzyme in eicosanoid production. cPLA2γ was reported to be located in endoplasmic reticulum (ER) and mitochondria and to have lysophospholipase activity beside phospholipase A2 (PLA2) activity. However, subcellular localization, mechanism of membrane binding, regulation and physiological function have not been fully established. In the present study, we examined the subcellular localization and enzymatic properties of cPLA2γ with C-terminal FLAG-tag. We found that cPLA2γ was located not only in ER but also mitochondria even in the absence of the prenylation. Purified recombinant cPLA2γ catalyzed an acyltransferase reaction from one molecule of lysophosphatidylcholine (LPC) to another, forming phosphatidylcholine (PC). LPC or lysophosphatidylethanolamine acted as acyl donor and acceptor, but lysophosphatidylserine, lysophosphatidylinositol and lysophosphatidic acid (LPA) did not. PC and phosphatidylethanolamine (PE) also acted as weak acyl donors. Reaction conditions changed the balance of lysophospholipase and Transacylation activities, with addition of LPA/PA, pH > 8, and elevated temperature markedly increasing Transacylation activity; this suggests that lysophospholipase/Transacylation activities of cPLA2γ may be regulated by various factors. As lysophospholipids are known to accumulate in ischemia heart and to induce arryhthmia, the cPLA2γ that is abundant in heart may have a protective role through clearance of lysophospholipids by its Transacylation activity.
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Subcellular localization and lysophospholipase/Transacylation activities of human group IVC phospholipase A2 (cPLA2γ)
Biochimica et biophysica acta, 2009Co-Authors: Atsushi Yamashita, Ken Tanaka, Ryo Kamata, Tsukasa Kumazawa, Naotaka Suzuki, Hiroki Koga, Keizo Waku, Takayuki SugiuraAbstract:Abstract cPLA2γ was identified as an ortholog of cPLA2α, which is a key enzyme in eicosanoid production. cPLA2γ was reported to be located in endoplasmic reticulum (ER) and mitochondria and to have lysophospholipase activity beside phospholipase A2 (PLA2) activity. However, subcellular localization, mechanism of membrane binding, regulation and physiological function have not been fully established. In the present study, we examined the subcellular localization and enzymatic properties of cPLA2γ with C-terminal FLAG-tag. We found that cPLA2γ was located not only in ER but also mitochondria even in the absence of the prenylation. Purified recombinant cPLA2γ catalyzed an acyltransferase reaction from one molecule of lysophosphatidylcholine (LPC) to another, forming phosphatidylcholine (PC). LPC or lysophosphatidylethanolamine acted as acyl donor and acceptor, but lysophosphatidylserine, lysophosphatidylinositol and lysophosphatidic acid (LPA) did not. PC and phosphatidylethanolamine (PE) also acted as weak acyl donors. Reaction conditions changed the balance of lysophospholipase and Transacylation activities, with addition of LPA/PA, pH > 8, and elevated temperature markedly increasing Transacylation activity; this suggests that lysophospholipase/Transacylation activities of cPLA2γ may be regulated by various factors. As lysophospholipids are known to accumulate in ischemia heart and to induce arryhthmia, the cPLA2γ that is abundant in heart may have a protective role through clearance of lysophospholipids by its Transacylation activity.
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Reverse reaction of lysophosphatidylinositol acyltransferase. Functional reconstitution of coenzyme A-dependent Transacylation system.
The Journal of biological chemistry, 2003Co-Authors: Atsushi Yamashita, Ryo Kamata, Masanobu Watanabe, Kazuaki Sato, Tomoyuki Miyashita, Tomonari Nagatsuka, Hironori Kondo, Norikazu Kawagishi, Hiroki Nakanishi, Takayuki SugiuraAbstract:CoA-dependent Transacylation activity in microsomes catalyzes the transfer of fatty acid between phospholipids and lysophospholipids in the presence of CoA without the generation of free fatty acid. We examined the mechanism of the Transacylation system using partially purified acyl-CoA:lysophosphatidylinositol (LPI) acyltransferase (LPIAT) from rat liver microsomes to test our hypothesis that both the reverse and forward reactions of acyl-CoA:lysophospholipid acyltransferases are involved in the CoA-dependent Transacylation process. The purified LPIAT fraction exhibited ATP-independent acyl-CoA synthetic activity and CoA-dependent LPI generation from PI, suggesting that LPIAT could operate in reverse to form acyl-CoA and LPI. CoA-dependent acylation of LPI by the purified LPIAT fraction required PI as the acyl donor. In addition, the combination of purified LPIAT and recombinant lysophosphatidic acid acyltransferase could reconstitute CoA-dependent Transacylation between PI and phosphatidic acid. These results suggest that the CoA-dependent Transacylation system consists of the following: 1) acyl-CoA synthesis from phospholipid through the reverse action of acyl-CoA:lysophospholipid acyltransferases; and 2) transfer of fatty acyl moiety from the newly formed acyl-CoA to lysophospholipid through the forward action of acyl-CoA:lysophospholipid acyltransferases.
Yang Xu - One of the best experts on this subject based on the ideXlab platform.
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the basis for acyl specificity in the tafazzin reaction
Journal of Biological Chemistry, 2017Co-Authors: Michael Schlame, Yang XuAbstract:: Tafazzin is a mitochondrial enzyme that transfers fatty acids from phospholipids to lysophospholipids. Mutations in tafazzin cause abnormal molecular species of cardiolipin and the clinical phenotype of Barth syndrome. However, the mechanism by which tafazzin creates acyl specificity has been controversial. We have shown that the lipid phase state can produce acyl specificity in the tafazzin reaction, but others have reported that tafazzin itself carries enzymatic specificity. To resolve this issue, we replicated and expanded the controversial experiments, i.e. the transfer of different acyl groups from phosphatidylcholine to monolysocardiolipin by yeast tafazzin. Our data show that this reaction requires the presence of detergent and does not take place in liposomes but in mixed micelles. To separate thermodynamic (lipid-dependent) from kinetic (enzyme-dependent) parameters, we followed the accumulation of cardiolipin during the reaction from the initial state to the equilibrium state. The Transacylation rates of different acyl groups varied over 2 orders of magnitude and correlated tightly with the concentration of cardiolipin in the equilibrium state (lipid-dependent parameter). In contrast, the rates by which different Transacylations approached the equilibrium state were very similar (enzyme-dependent parameter). Furthermore, we found that tafazzin catalyzes the remodeling of cardiolipin by combinations of forward and reverse Transacylations, essentially creating an equilibrium distribution of acyl groups. These data strongly support the idea that the acyl specificity of the tafazzin reaction results from the physical properties of lipids.
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reconstitution of acyl specific phospholipid remodeling by purified tafazzin in vitro
Biophysical Journal, 2012Co-Authors: Michael Schlame, Bob Berno, Devrim Acehan, Yang Xu, David L Stokes, Richard M EpandAbstract:Cardiolipin is a mitochondrial phospholipid with a unique composition and distribution of acyl groups. The cardiolipin composition depends on tafazzin, a phospholipid-lysophospholipid transacylase, although the enzyme itself lacks acyl specificity. We incubated isolated tafazzin with various mixtures of phospholipids and lysophospholipids, characterized the lipid phase state by 31P-NMR, and measured newly formed molecular species by mass spectrometry. Significant Transacylation activity was observed only in non-bilayer lipid aggregates, in which lipids had a low packing order. The lipid phase state profoundly affected the substrate specificity of the tafazzin reaction. In particular, tetralinoleoyl-cardiolipin, a prototype molecular species, formed only under conditions that favor the inverted hexagonal phase. In isolated mitochondria, less than 2 percent of lipids participated in Transacylations, suggesting that tafazzin acts only on privileged lipid domains. We propose that tafazzin reacts with non-bilayer lipids in mitochondria and that acyl specificity arises from spontaneous self-organization of these domains.View Large Image | View Hi-Res Image | Download PowerPoint Slide
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the enzymatic function of tafazzin
Journal of Biological Chemistry, 2006Co-Authors: Yang Xu, Ashim Malhotra, Michael SchlameAbstract:Abstract Tafazzin is a putative enzyme that is involved in cardiolipin metabolism, it may carry mutations responsible for Barth syndrome. To identify the biochemical reaction catalyzed by tafazzin, we expressed the full-length isoform of Drosophila melanogaster tafazzin in a baculovirus-Sf9 insect cell system. Tafazzin expression induced a new enzymatic function in Sf9 cell mitochondria, namely 1-palmitoyl-2-[14C]linoleoyl-phosphatidylcholine:monolysocardiolipin linoleoyltransferase. We also found evidence for the reverse reaction, because tafazzin expression caused transfer of acyl groups from phospholipids to 1-[14C]palmitoyl-2-lyso-phosphatidylcholine. An affinity-purified tafazzin construct, tagged with the maltose-binding protein, catalyzed both forward and reverse Transacylations between cardiolipin and phosphatidylcholine, but was unable to utilize CoA or acyl-CoA as substrates. Whereas tafazzin supported Transacylations between various phospholipid-lysophospholipid pairs, it showed the highest rate for the phosphatidylcholine-cardiolipin Transacylation. Transacylation activities were about 10-fold higher for linoleoyl groups than for oleoyl groups, and they were negligible for arachidonoyl groups. The data show that Drosophila tafazzin is a CoA-independent, acyl-specific phospholipid transacylase with substrate preference for cardiolipin and phosphatidylcholine.
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Remodeling of Cardiolipin by Phospholipid Transacylation
Journal of Biological Chemistry, 2003Co-Authors: Yang Xu, Richard I. Kelley, Thomas J J Blanck, Michael SchlameAbstract:Abstract Mitochondrial cardiolipin (CL) contains unique fatty acid patterns, but it is not known how the characteristic molecular species of CL are formed. We found a novel reaction that transfers acyl groups from phosphatidylcholine or phosphatidylethanolamine to CL in mitochondria of rat liver and human lymphoblasts. Acyl transfer was stimulated by ADP, ATP, and ATPγS, but not by other nucleotides. Coenzyme A stimulated the reaction only in the absence of adenine nucleotides. Free fatty acids were not incorporated into CL under the same incubation condition. The Transacylation required addition of exogenous CL or monolyso-CL, whereas dilyso-CL was not a substrate. Transacylase activity was decreased in lymphoblasts from patients with Barth syndrome (tafazzin deletion), and this was accompanied by drastic changes in the molecular composition of CL. In rat liver, where linoleic acid was the most abundant residue of CL, only linoleoyl groups were transferred into CL, but not oleoyl or arachidonoyl groups. We demonstrated complete remodeling of tetraoleoyl-CL to tetralinoleoyl-CL in rat liver mitochondria and identified the intermediates linoleoyl-trioleoyl-CL, dilinoleoyl-dioleoyl-CL, and trilinoleoyl-oleoyl-CL by high-performance liquid chromatography. The data suggest that CL is remodeled by acyl specific phospholipid Transacylation and that tafazzin is an acyltransferase involved in this mechanism.
Keizo Waku - One of the best experts on this subject based on the ideXlab platform.
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acyltransferases and transacylases that determine the fatty acid composition of glycerolipids and the metabolism of bioactive lipid mediators in mammalian cells and model organisms
Progress in Lipid Research, 2014Co-Authors: Atsushi Yamashita, Keizo Waku, Yasuhiro Hayashi, Saori Oka, Takashi Tanikawa, Yoko Nemotosasaki, Makoto Ito, Takayuki SugiuraAbstract:Abstract Over one hundred different phospholipid molecular species are known to be present in mammalian cells and tissues. Fatty acid remodeling systems for phospholipids including acyl-CoA:lysophospholipid acyltransferases, CoA-dependent and CoA-independent Transacylation systems, are involved in the biosynthesis of these molecular species. Acyl-CoA:lysophospholipid acyltransferase system is involved in the synthesis of phospholipid molecular species containing sn-1 saturated and sn-2 unsaturated fatty acids. The CoA-dependent Transacylation system catalyzes the transfer of fatty acids esterified in phospholipids to lysophospholipids in the presence of CoA without the generation of free fatty acids. The CoA-dependent Transacylation reaction in the rat liver exhibits strict fatty acid specificity, i.e., three types of fatty acids (20:4, 18:2 and 18:0) are transferred. On the other hand, CoA-independent transacylase catalyzes the transfer of C20 and C22 polyunsaturated fatty acids from diacyl phospholipids to various lysophospholipids, especially ether-containing lysophospholipids, in the absence of any cofactors. CoA-independent transacylase is assumed to be involved in the accumulation of PUFA in ether-containing phospholipids. These enzymes are involved in not only the remodeling of fatty acids, but also the synthesis and degradation of some bioactive lipids and their precursors. In this review, recent progresses in acyltransferase research including the identification of the enzyme’s genes are described.
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subcellular localization and lysophospholipase Transacylation activities of human group ivc phospholipase a2 cpla2γ
Biochimica et Biophysica Acta, 2009Co-Authors: Atsushi Yamashita, Ken Tanaka, Ryo Kamata, Tsukasa Kumazawa, Naotaka Suzuki, Hiroki Koga, Keizo Waku, Takayuki SugiuraAbstract:Abstract cPLA2γ was identified as an ortholog of cPLA2α, which is a key enzyme in eicosanoid production. cPLA2γ was reported to be located in endoplasmic reticulum (ER) and mitochondria and to have lysophospholipase activity beside phospholipase A2 (PLA2) activity. However, subcellular localization, mechanism of membrane binding, regulation and physiological function have not been fully established. In the present study, we examined the subcellular localization and enzymatic properties of cPLA2γ with C-terminal FLAG-tag. We found that cPLA2γ was located not only in ER but also mitochondria even in the absence of the prenylation. Purified recombinant cPLA2γ catalyzed an acyltransferase reaction from one molecule of lysophosphatidylcholine (LPC) to another, forming phosphatidylcholine (PC). LPC or lysophosphatidylethanolamine acted as acyl donor and acceptor, but lysophosphatidylserine, lysophosphatidylinositol and lysophosphatidic acid (LPA) did not. PC and phosphatidylethanolamine (PE) also acted as weak acyl donors. Reaction conditions changed the balance of lysophospholipase and Transacylation activities, with addition of LPA/PA, pH > 8, and elevated temperature markedly increasing Transacylation activity; this suggests that lysophospholipase/Transacylation activities of cPLA2γ may be regulated by various factors. As lysophospholipids are known to accumulate in ischemia heart and to induce arryhthmia, the cPLA2γ that is abundant in heart may have a protective role through clearance of lysophospholipids by its Transacylation activity.
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Subcellular localization and lysophospholipase/Transacylation activities of human group IVC phospholipase A2 (cPLA2γ)
Biochimica et biophysica acta, 2009Co-Authors: Atsushi Yamashita, Ken Tanaka, Ryo Kamata, Tsukasa Kumazawa, Naotaka Suzuki, Hiroki Koga, Keizo Waku, Takayuki SugiuraAbstract:Abstract cPLA2γ was identified as an ortholog of cPLA2α, which is a key enzyme in eicosanoid production. cPLA2γ was reported to be located in endoplasmic reticulum (ER) and mitochondria and to have lysophospholipase activity beside phospholipase A2 (PLA2) activity. However, subcellular localization, mechanism of membrane binding, regulation and physiological function have not been fully established. In the present study, we examined the subcellular localization and enzymatic properties of cPLA2γ with C-terminal FLAG-tag. We found that cPLA2γ was located not only in ER but also mitochondria even in the absence of the prenylation. Purified recombinant cPLA2γ catalyzed an acyltransferase reaction from one molecule of lysophosphatidylcholine (LPC) to another, forming phosphatidylcholine (PC). LPC or lysophosphatidylethanolamine acted as acyl donor and acceptor, but lysophosphatidylserine, lysophosphatidylinositol and lysophosphatidic acid (LPA) did not. PC and phosphatidylethanolamine (PE) also acted as weak acyl donors. Reaction conditions changed the balance of lysophospholipase and Transacylation activities, with addition of LPA/PA, pH > 8, and elevated temperature markedly increasing Transacylation activity; this suggests that lysophospholipase/Transacylation activities of cPLA2γ may be regulated by various factors. As lysophospholipids are known to accumulate in ischemia heart and to induce arryhthmia, the cPLA2γ that is abundant in heart may have a protective role through clearance of lysophospholipids by its Transacylation activity.
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ATP-independent Fatty Acyl-Coenzyme A Synthesis from Phospholipid COENZYME A-DEPENDENT Transacylation ACTIVITY TOWARD LYSOPHOSPHATIDIC ACID CATALYZED BY ACYL-COENZYME A:LYSOPHOSPHATIDIC ACID ACYLTRANSFERASE
The Journal of biological chemistry, 2001Co-Authors: Atsushi Yamashita, Takayuki Sugiura, Tomoyuki Miyashita, Tomonari Nagatsuka, Norikazu Kawagishi, Kazuhiko Kume, Takao Shimizu, Keizo WakuAbstract:Abstract CoA-dependent Transacylation activity in microsomes is known to catalyze the transfer of fatty acids between phospholipids and lysophospholipids in the presence of CoA without the generation of free fatty acids. We previously found a novel acyl-CoA synthetic pathway, ATP-independent acyl-CoA synthesis from phospholipids. We proposed that: 1) the ATP-independent acyl-CoA synthesis is due to the reverse reaction of acyl-CoA:lysophospholipid acyltransferases and 2) the reverse and forward reactions of acyltransferases can combine to form a CoA-dependent Transacylation system. To test these proposals, we examined whether or not recombinant mouse acyl-CoA:1-acyl-sn-glycero-3-phosphate (lysophosphatidic acid, LPA) acyltransferase (LPAAT) could catalyze ATP-independent acyl-CoA synthetic activity and CoA-dependent Transacylation activity. ATP-independent acyl-CoA synthesis was indeed found in the membrane fraction from Escherichia coli cells expressing mouse LPAAT, whereas negligible activity was observed in mock-transfected cells. Phosphatidic acid (PA), but not free fatty acids, served as an acyl donor for the reaction, and LPA was formed from PA in a CoA-dependent manner during acyl-CoA synthesis. These results indicate that the ATP-independent acyl-CoA synthesis was due to the reverse reaction of LPAAT. In addition, bacterial membranes containing LPAAT catalyzed CoA-dependent acylation of LPA; PA but not free fatty acid served as an acyl donor. These results indicate that the CoA-dependent Transacylation of LPA consists of 1) acyl-CoA synthesis from PA through the reverse action of LPAAT and 2) the transfer of the fatty acyl moiety of the newly formed acyl-CoA to LPA through the forward reaction of LPAAT.
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Induction of coenzyme A-dependent Transacylation activity in rat liver microsomes by administration of clofibrate
Biochimica et biophysica acta, 1994Co-Authors: Atsushi Yamashita, Takayuki Sugiura, Masanobu Watanabe, Kazuaki Sato, Yoshihiro Tokudome, Keizo WakuAbstract:Abstract The effect of administration of clofibrate on the activity of coenzyme A-dependent (CoA-dependent) Transacylation of 1-acyl-glycerophosphocholine (1-acyl-GPC) was examined in rat liver microsomes. Administration of clofibrate to rats increased the activity of CoA-dependent Transacylation of 1-[14C]acyl-GPC and the activity reached a value (8.37 nmol/min per mg protein) twice that in control rats (3.95 nmol/min per mg protein) without any changes in apparent Km values for CoA (1.2 μM in control and 1.0 μM in clofibrate-treated) and 1-acyl-GPC (33.4 μM in control and 27.8 μM in clofibrate-treated). The rate of CoA-dependent transfer of [14C]arachidonic acid (20:4) from 1-acyl-2-[14C]20:4-glycerophosphoethanolamine (GPE) or 1-acyl-2-[14C]20:4-glycerophosphoinositol (GPI) to 1-acyl-GPC (synthesis of 1-acyl-2-[14C]20:4-GPC) was also increased by treatment with clofibrate (1.9-fold and 1.5-fold increases, respectively). These results suggest that a CoA-dependent Transacylation system of 1-acyl-GPC was induced by treatment with clofibrate.
