PMID:22753059

Citation	Yuan, Y, Sachdeva, M, Leeds, JA and Meredith, TC (2012) Fatty acid biosynthesis in Pseudomonas aeruginosa is initiated by the FabY class of β-ketoacyl acyl carrier protein synthases. J. Bacteriol. 194:5171-84
Abstract	The prototypical type II fatty acid synthesis (FAS) pathway in bacteria utilizes two distinct classes of β-ketoacyl synthase (KAS) domains to assemble long-chain fatty acids, the KASIII domain for initiation and the KASI/II domain for elongation. The central role of FAS in bacterial viability and virulence has stimulated significant effort toward developing KAS inhibitors, particularly against the KASIII domain of the β-acetoacetyl-acyl carrier protein (ACP) synthase FabH. Herein, we show that the opportunistic pathogen Pseudomonas aeruginosa does not utilize a FabH ortholog but rather a new class of divergent KAS I/II enzymes to initiate the FAS pathway. When a P. aeruginosa cosmid library was used to rescue growth in a fabH downregulated strain of Escherichia coli, a single unannotated open reading frame, PA5174, complemented fabH depletion. While deletion of all four KASIII domain-encoding genes in the same P. aeruginosa strain resulted in a wild-type growth phenotype, deletion of PA5174 alone specifically attenuated growth due to a defect in de novo FAS. Siderophore secretion and quorum-sensing signaling, particularly in the rhl and Pseudomonas quinolone signal (PQS) systems, was significantly muted in the absence of PA5174. The defect could be repaired by intergeneric complementation with E. coli fabH. Characterization of recombinant PA5174 confirmed a preference for short-chain acyl coenzyme A (acyl-CoA) substrates, supporting the identification of PA5174 as the predominant enzyme catalyzing the condensation of acetyl coenzyme A with malonyl-ACP in P. aeruginosa. The identification of the functional role for PA5174 in FAS defines the new FabY class of β-ketoacyl synthase KASI/II domain condensation enzymes.
Links	PubMed PMC3457228 Online version:10.1128/JB.00792-12
Keywords	3-Oxoacyl-(Acyl-Carrier-Protein) Synthase/classification; 3-Oxoacyl-(Acyl-Carrier-Protein) Synthase/metabolism; Amino Acid Sequence; Bacterial Proteins/genetics; Bacterial Proteins/metabolism; Binding Sites; Down-Regulation; Fatty Acids/biosynthesis; Gene Deletion; Gene Expression Regulation, Bacterial; Gene Expression Regulation, Enzymologic; Phylogeny; Protein Structure, Tertiary; Pseudomonas aeruginosa/enzymology; Pseudomonas aeruginosa/genetics; Pseudomonas aeruginosa/metabolism
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Annotations

Gene product	Qualifier	GO Term	Evidence Code	with/from	Aspect	Extension	Notes	Status
PSEAE:FABY		GO:0033818: beta-ketoacyl-acyl-carrier-protein synthase III activity	ECO:0000314:		F		Figure 8a: NADPH consumption of PA5174 and E. coli FabH were similar	complete CACAO 9288
PSEAE:FABY		GO:0006633: fatty acid biosynthetic process	ECO:0000315:		P		Figure 4B: The PA5174 incapacitated mutant displayed a lower percent incorporated label than the wild type.	complete CACAO 9486
PSEAE:FABY		GO:0009372: quorum sensing	ECO:0000315:		P		Figure 5A: Production of acyl-homoserine lactone decreased in PA5174 incapacitated mutants and increased in PA5174 complemented mutants (TMT41) Figure 5B: Wild-type strain and TMT41 mutants produced halos indicating ion-pairing complexation between methylene blue dye, cetyltrimethylammonium bromide, and secreted rhamnolipids but mutant with nonfunctional PA5174 did not. Figure 5C: Rhamnose levels were higher in TMT41 mutant than in wild type. Rhamnose levels were also lower in mutant with nonfunctional PA5174 than in wild type.	complete CACAO 9492
PSEAE:FABY		GO:0019290: siderophore biosynthetic process	ECO:0000315:		P		Figure 7: Wild-type, mutant strain without PA5174, PA5174 back-complemented with plasmid, and FabH-complemented strain were streaked on LB-chrome azurol S indicator plates. Only the PA5174 deficient strain displayed no signs of iron sequestration (no yellow border around bacteria).	complete CACAO 9514
PSEAE:FABY	enables	GO:0033818: beta-ketoacyl-acyl-carrier-protein synthase III activity	ECO:0000314: direct assay evidence used in manual assertion		F		Seeded From UniProt	complete
PSEAE:FABY	involved_in	GO:0006633: fatty acid biosynthetic process	ECO:0000315: mutant phenotype evidence used in manual assertion		P		Seeded From UniProt	complete
PSEAE:FABY	involved_in	GO:0009372: quorum sensing	ECO:0000315: mutant phenotype evidence used in manual assertion		P		Seeded From UniProt	complete
PSEAE:FABY	involved_in	GO:0019290: siderophore biosynthetic process	ECO:0000315: mutant phenotype evidence used in manual assertion		P		Seeded From UniProt	complete

References

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