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Ralstonia solanacearum RSp0194 Encodes a Novel 3-Keto-Acyl Carrier Protein Synthase III.

Mao YH, Ma JC, Li F, Hu Z, Wang HH - PLoS ONE (2015)

Bottom Line: Although the RsfabW mutant was viable, RsfabW was responsible for RsfabH mutant growth on medium containing free fatty acids.Our results also showed that RsFabW could condense acyl-ACP (C4-ACP to C8-ACP) with malonyl-ACP, to produce 3-keto-acyl-ACP in vitro, which implies that RsFabW plays a special role in fatty acid synthesis of R. solanacearum.All of these data confirm that R. solanacearum not only utilizes acetyl-CoA, but also, utilizes medium-chain acyl-CoAs or acyl-ACPs as primers to initiate fatty acid synthesis.

View Article: PubMed Central - PubMed

Affiliation: Guangdong Provincial Key Laboratory of Protein Function and Regulation in Agricultural Organisms, College of Life Sciences, South China Agricultural University, Guangzhou, Guangdong 510642, China.

ABSTRACT
Fatty acid synthesis (FAS), a primary metabolic pathway, is essential for survival of bacteria. Ralstonia solanacearum, a β-proteobacteria member, causes a bacterial wilt affecting more than 200 plant species, including many economically important plants. However, thus far, the fatty acid biosynthesis pathway of R. solanacearum has not been well studied. In this study, we characterized two forms of 3-keto-ACP synthase III, RsFabH and RsFabW, in R. solanacearum. RsFabH, the homologue of Escherichia coli FabH, encoded by the chromosomal RSc1050 gene, catalyzes the condensation of acetyl-CoA with malonyl-ACP in the initiation steps of fatty acid biosynthesis in vitro. The RsfabH mutant lost de novo fatty acid synthetic ability, and grows in medium containing free fatty acids. RsFabW, a homologue of Pseudomonas aeruginosa PA3286, encoded by a megaplasmid gene, RSp0194, condenses acyl-CoA (C2-CoA to C10-CoA) with malonyl-ACP to produce 3-keto-acyl-ACP in vitro. Although the RsfabW mutant was viable, RsfabW was responsible for RsfabH mutant growth on medium containing free fatty acids. Our results also showed that RsFabW could condense acyl-ACP (C4-ACP to C8-ACP) with malonyl-ACP, to produce 3-keto-acyl-ACP in vitro, which implies that RsFabW plays a special role in fatty acid synthesis of R. solanacearum. All of these data confirm that R. solanacearum not only utilizes acetyl-CoA, but also, utilizes medium-chain acyl-CoAs or acyl-ACPs as primers to initiate fatty acid synthesis.

No MeSH data available.


Related in: MedlinePlus

Model of the fatty acid biosynthesis pathway in R. solanacearum.Abbreviations: Acc, acetyl-CoA carboxylase; FabD, malonyl-CoA: ACP transacylase; RsFabH, 3-ketoacyl-ACP synthase III; RsFabW, 3-ketoacyl-ACP synthase III; FabG, 3-ketoacyl-ACP reductase; FabZ, 3-hydroxyacyl-ACP dehydratase; RsFabF, 3-ketoacyl-ACP synthase II; FabI, enoyl-ACP reductase. The full line indicates RsFabH uses acetyl-CoA substrate to initiate the fatty acid synthesis. The dotted line indicates RsFabW uses acyl-CoAs as substrates to initiate the fatty acid synthesis. The dashed line indicates RsFabW uses acyl-ACPs as substrates to initiate the fatty acid synthesis.
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pone.0136261.g006: Model of the fatty acid biosynthesis pathway in R. solanacearum.Abbreviations: Acc, acetyl-CoA carboxylase; FabD, malonyl-CoA: ACP transacylase; RsFabH, 3-ketoacyl-ACP synthase III; RsFabW, 3-ketoacyl-ACP synthase III; FabG, 3-ketoacyl-ACP reductase; FabZ, 3-hydroxyacyl-ACP dehydratase; RsFabF, 3-ketoacyl-ACP synthase II; FabI, enoyl-ACP reductase. The full line indicates RsFabH uses acetyl-CoA substrate to initiate the fatty acid synthesis. The dotted line indicates RsFabW uses acyl-CoAs as substrates to initiate the fatty acid synthesis. The dashed line indicates RsFabW uses acyl-ACPs as substrates to initiate the fatty acid synthesis.

Mentions: The growth of RsfabH mutants on medium containing long-chain fatty acids (C14 to C18) was weaker than on medium in the presence of medium-chain fatty acids (C8 to C12) (Fig 3). This was in agreement with the substrate specificity of RsFabW for C8-CoA and C10-CoA in vitro (Table 1). This indicated that exogenous long-chain fatty acids (C14 to C18) should be degraded to C8-CoA by β-oxidation, and then RsFabW would shunt C8-CoA into the fatty acid synthesis pathway to make LPS, UFA, and SFA required for bacteria growth. However, although FabW possessed strong sequence similarity to P. aeruginosa PA3286 (Fig 1D), FabW was different from P. aeruginosa PA3286 [9]. First, FabW not only uses acyl-CoAs (such as octanoyl-CoA or decanoyl-CoA) as primers, but also condenses short-chain acyl-ACPs (such as butanoyl-ACP, hexanoyl-ACP, and octanoyl-ACP) with malonyl-ACP. Second, FabW seems to have a unique role in fatty acid synthesis in R. solanacearum, but does not only shunt intermediates from β-oxidation degradation into fatty acid biosynthesis. R. solanacearum mainly invades xylem vessels of host plants [35], where this is a lack of sufficient phospholipids or free fatty acids to support this bacterial growth. RsFabW is also distinct from long-chain 3-ketoacyl-ACP synthase I/II, which can catalyze the condensation of long-chain acyl-ACP with malonyl-ACP in the elongation cycle of bacterial fatty acid synthesis [10]. RsFabW did not utilize decanoyl-ACP or longer acyl-ACPs as substrates. It has been demonstrated that R. solanacearum only has one long chain 3-ketoacyl-acyl carrier protein synthase, RsFabF1, which possesses both the activity of 3-ketoacyl-ACP synthase II and I [23]. Therefore, it is possible that RsFabW helps RsFabF1 to function in elongation reactions of medium-chain fatty acid synthesis. On the basis of our results, we suggest a model for fatty acid biosynthesis in R. solanacearum in Fig 6.


Ralstonia solanacearum RSp0194 Encodes a Novel 3-Keto-Acyl Carrier Protein Synthase III.

Mao YH, Ma JC, Li F, Hu Z, Wang HH - PLoS ONE (2015)

Model of the fatty acid biosynthesis pathway in R. solanacearum.Abbreviations: Acc, acetyl-CoA carboxylase; FabD, malonyl-CoA: ACP transacylase; RsFabH, 3-ketoacyl-ACP synthase III; RsFabW, 3-ketoacyl-ACP synthase III; FabG, 3-ketoacyl-ACP reductase; FabZ, 3-hydroxyacyl-ACP dehydratase; RsFabF, 3-ketoacyl-ACP synthase II; FabI, enoyl-ACP reductase. The full line indicates RsFabH uses acetyl-CoA substrate to initiate the fatty acid synthesis. The dotted line indicates RsFabW uses acyl-CoAs as substrates to initiate the fatty acid synthesis. The dashed line indicates RsFabW uses acyl-ACPs as substrates to initiate the fatty acid synthesis.
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pone.0136261.g006: Model of the fatty acid biosynthesis pathway in R. solanacearum.Abbreviations: Acc, acetyl-CoA carboxylase; FabD, malonyl-CoA: ACP transacylase; RsFabH, 3-ketoacyl-ACP synthase III; RsFabW, 3-ketoacyl-ACP synthase III; FabG, 3-ketoacyl-ACP reductase; FabZ, 3-hydroxyacyl-ACP dehydratase; RsFabF, 3-ketoacyl-ACP synthase II; FabI, enoyl-ACP reductase. The full line indicates RsFabH uses acetyl-CoA substrate to initiate the fatty acid synthesis. The dotted line indicates RsFabW uses acyl-CoAs as substrates to initiate the fatty acid synthesis. The dashed line indicates RsFabW uses acyl-ACPs as substrates to initiate the fatty acid synthesis.
Mentions: The growth of RsfabH mutants on medium containing long-chain fatty acids (C14 to C18) was weaker than on medium in the presence of medium-chain fatty acids (C8 to C12) (Fig 3). This was in agreement with the substrate specificity of RsFabW for C8-CoA and C10-CoA in vitro (Table 1). This indicated that exogenous long-chain fatty acids (C14 to C18) should be degraded to C8-CoA by β-oxidation, and then RsFabW would shunt C8-CoA into the fatty acid synthesis pathway to make LPS, UFA, and SFA required for bacteria growth. However, although FabW possessed strong sequence similarity to P. aeruginosa PA3286 (Fig 1D), FabW was different from P. aeruginosa PA3286 [9]. First, FabW not only uses acyl-CoAs (such as octanoyl-CoA or decanoyl-CoA) as primers, but also condenses short-chain acyl-ACPs (such as butanoyl-ACP, hexanoyl-ACP, and octanoyl-ACP) with malonyl-ACP. Second, FabW seems to have a unique role in fatty acid synthesis in R. solanacearum, but does not only shunt intermediates from β-oxidation degradation into fatty acid biosynthesis. R. solanacearum mainly invades xylem vessels of host plants [35], where this is a lack of sufficient phospholipids or free fatty acids to support this bacterial growth. RsFabW is also distinct from long-chain 3-ketoacyl-ACP synthase I/II, which can catalyze the condensation of long-chain acyl-ACP with malonyl-ACP in the elongation cycle of bacterial fatty acid synthesis [10]. RsFabW did not utilize decanoyl-ACP or longer acyl-ACPs as substrates. It has been demonstrated that R. solanacearum only has one long chain 3-ketoacyl-acyl carrier protein synthase, RsFabF1, which possesses both the activity of 3-ketoacyl-ACP synthase II and I [23]. Therefore, it is possible that RsFabW helps RsFabF1 to function in elongation reactions of medium-chain fatty acid synthesis. On the basis of our results, we suggest a model for fatty acid biosynthesis in R. solanacearum in Fig 6.

Bottom Line: Although the RsfabW mutant was viable, RsfabW was responsible for RsfabH mutant growth on medium containing free fatty acids.Our results also showed that RsFabW could condense acyl-ACP (C4-ACP to C8-ACP) with malonyl-ACP, to produce 3-keto-acyl-ACP in vitro, which implies that RsFabW plays a special role in fatty acid synthesis of R. solanacearum.All of these data confirm that R. solanacearum not only utilizes acetyl-CoA, but also, utilizes medium-chain acyl-CoAs or acyl-ACPs as primers to initiate fatty acid synthesis.

View Article: PubMed Central - PubMed

Affiliation: Guangdong Provincial Key Laboratory of Protein Function and Regulation in Agricultural Organisms, College of Life Sciences, South China Agricultural University, Guangzhou, Guangdong 510642, China.

ABSTRACT
Fatty acid synthesis (FAS), a primary metabolic pathway, is essential for survival of bacteria. Ralstonia solanacearum, a β-proteobacteria member, causes a bacterial wilt affecting more than 200 plant species, including many economically important plants. However, thus far, the fatty acid biosynthesis pathway of R. solanacearum has not been well studied. In this study, we characterized two forms of 3-keto-ACP synthase III, RsFabH and RsFabW, in R. solanacearum. RsFabH, the homologue of Escherichia coli FabH, encoded by the chromosomal RSc1050 gene, catalyzes the condensation of acetyl-CoA with malonyl-ACP in the initiation steps of fatty acid biosynthesis in vitro. The RsfabH mutant lost de novo fatty acid synthetic ability, and grows in medium containing free fatty acids. RsFabW, a homologue of Pseudomonas aeruginosa PA3286, encoded by a megaplasmid gene, RSp0194, condenses acyl-CoA (C2-CoA to C10-CoA) with malonyl-ACP to produce 3-keto-acyl-ACP in vitro. Although the RsfabW mutant was viable, RsfabW was responsible for RsfabH mutant growth on medium containing free fatty acids. Our results also showed that RsFabW could condense acyl-ACP (C4-ACP to C8-ACP) with malonyl-ACP, to produce 3-keto-acyl-ACP in vitro, which implies that RsFabW plays a special role in fatty acid synthesis of R. solanacearum. All of these data confirm that R. solanacearum not only utilizes acetyl-CoA, but also, utilizes medium-chain acyl-CoAs or acyl-ACPs as primers to initiate fatty acid synthesis.

No MeSH data available.


Related in: MedlinePlus