Defining a Structural and Kinetic Rationale for Paralogous Copies of Phenylacetate-CoA Ligases from the Cystic Fibrosis Pathogen Burkholderia cenocepacia J2315

Law, Adrienne; Boulanger, Martin J.

doi:10.1074/jbc.m111.219683

Cited by 30 publications

(41 citation statements)

References 36 publications

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“…The paa gene clusters that encode the enzymes responsible for PA catabolism are widely distributed in bacteria, and they are present in about 16% of completely sequenced bacterial genomes and in some archaea (Ferrández et al ., ; Olivera et al ., ; Mohamed et al ., ; Rost et al ., ; Alonso et al ., ; Bartolome‐Martín et al ., ; Kunishima et al ., ; Navarro‐Llorens et al ., 2005; 2008; Song et al ., ; Di Gennaro et al ., ; Hamlin et al ., ; Teufel et al ., ; Law and Boulanger, ; Patrauchan et al ., ; Sakamoto et al ., ). The paa pathway is controlled by a transcriptional repressor named PaaX or PaaR depending on the microorganism (Ferrández et al ., ; Olivera et al ., ; Navarro‐Llorens et al ., ; Hamlin et al ., ).…”

Section: Introductionmentioning

confidence: 99%

Insights on the regulation of the phenylacetate degradation pathway from Escherichia coli

Fernández

Dı́az

Garcı́a

2013

Environ Microbiol Rep

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The paa genes for phenylacetic acid (PA) catabolism encode the best characterized aerobic hybrid route involved in the bacterial degradation of aromatic compounds. Here, we demonstrate that the divergent paaZ and paaA-K catabolic operons of Escherichia coli are regulated by two genes, paaXY, that form a distinct transcriptional unit driven by the Px promoter. In vivo and in vitro approaches using purified PaaX regulatory protein revealed that this regulator is able to bind and inhibit the activity of Px in a phenylacetyl-coenzyme A (PA-CoA) dependent manner. The autoregulation of paaXY is due to the competition between PaaX and RNA polymerase for binding to the regulatory Px promoter. Whereas a similar mechanism of repression mediated by PaaX was shown to occur at the catabolic Pz promoter; the catabolic Pa promoter is inhibited by PaaX by a mechanism that does not involves competition with RNA polymerase. We have shown for the first time that the paaY gene product is essential for an efficient growth in PA. Purified PaaY was shown to be a trimer in solution with a broad thioesterase activity stimulated by some metals. This thioesterase activity will allow the detoxification of some CoA-intermediates that block the aerobic catabolism of PA, as previously suggested, but also will avoid the accumulation of some CoA derivatives that could behave as antagonists of the inducer effect caused by PA-CoA on the PaaX repressor for an efficient expression of the paa genes. This regulatory function mediated by PaaY constitutes an additional regulatory checkpoint that makes the circuit that controls the transcription of the paa genes more complex than previously thought, and it could represent a general strategy present in most bacterial paa gene clusters that also harbour the paaY gene.

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Section: Introductionmentioning

confidence: 99%

Insights on the regulation of the phenylacetate degradation pathway from Escherichia coli

Fernández

Dı́az

Garcı́a

2013

Environ Microbiol Rep

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“…13,18,20,21 It has been hypothesized that in different CoA ligase enzymes, there is an equilibrium between conformations 1 and 2, and this equilibrium may be different for different CoA ligases during crystallization. 17 In the AuaEII crystal structure, the C-terminal domains of both monomers have elevated B-factors for the majority of residues except in the A10 core region, where Lys506 interacts with the anthraniloyl-AMP intermediate ( Figure S3).…”

mentioning

confidence: 99%

“…To date, many CoA ligase crystal structures have been solved that are bound with an acid substrate or aryl-adenylate intermediate, and these structures reveal residues that are important protein−substrate interactions. 13,17,18,21 An in depth comparison of the AuaEII crystal structure to the energy minimized AuaE homology model and six previously determined aryl:CoA ligase crystal structures reveal unprecedented insights into aryl substrate binding ( Figure 3; Table 1). The aryl-binding region of AuaEII consists of histidine (His324), isoleucine (Ile325), alanine (Ala293), and phenyalanine (Phe222).…”

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confidence: 99%

Structural Insights into Anthranilate Priming during Type II Polyketide Biosynthesis

Jackson¹,

Tu²,

Nguyen³

et al. 2015

ACS Chem. Biol.

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The incorporation of nonacetate starter units during type II polyketide biosynthesis helps diversify natural products. Currently, there are few enzymatic strategies for the incorporation of nonacetate starter units in type II polyketide synthase (PKS) pathways. Here we report the crystal structure of AuaEII, the anthranilate:CoA ligase responsible for the generation of anthraniloyl-CoA, which is used as a starter unit by a type II PKS in aurachin biosynthesis. We present structural and protein sequence comparisons to other aryl:CoA ligases. We also compare the AuaEII crystal structure to a model of a CoA ligase homologue, AuaE, which is present in the same gene cluster. AuaE is predicted to have the same fold as AuaEII, but instead of CoA ligation, AuaE catalyzes acyl transfer of anthranilate from anthraniloyl-CoA to the acyl carrier protein (ACP). Together, this work provides insight into the molecular basis for starter unit selection of anthranilate in type II PKS biosynthesis.

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“…According to this finding, Law and Boulanger (2011) suggested that the more active, and substrate-versatile PaaK1 might be more beneficial during infection. However, our results show that PaaK1 (and possibly 3-OHPA as a PA degradation inducer) does not play a significant role in pathogenicity.…”

Section: Discussionmentioning

confidence: 94%

“…Given the evidence that 3-OHPA and 4-OHPA can be substrates of the PaaK enzymes (Law and Boulanger, 2011) we also considered that these compounds could be funneled exclusively through the PA catabolic pathway. B. cenocepacia K56-2 was grown in 3-OHPA or 4-OHPA as sole carbon sources and growth was recorded as final optical density.…”

Section: Resultsmentioning

confidence: 99%

3-Hydroxyphenylacetic Acid Induces the Burkholderia cenocepacia Phenylacetic Acid Degradation Pathway ? Toward Understanding the Contribution of Aromatic Catabolism to Pathogenesis

Imolorhe¹,

Cardona²

2011

Front. Cell. Inf. Microbio.

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The phenylacetic acid (PA) degradative pathway is the central pathway by which various aromatic compounds (e.g., styrene) are degraded. Upper pathways for different aromatic compounds converge at common intermediate phenylacetyl-CoA (PA-CoA), which is then metabolized to succinyl-CoA and acetyl-CoA. We previously made a link in Burkholderia cenocepacia between PA degradation and virulence by showing that insertional mutagenesis of paaA and paaE genes, that encode part of a multicomponent oxidase of PA-CoA, results in PA-conditional growth and an attenuated killing phenotype in the Caenorhabditis elegans model of infection. However, insertional mutagenesis of paaK1, which encodes a phenylacetate-CoA ligase, did not result in a PA-conditional growth probably due to the presence of a putative paralog gene paaK2. Recently published crystallographic and enzyme kinetics data comparing the two PaaK ligases showed that PaaK1 is more active than PaaK2 and that the larger binding pocket of PaaK1 can accommodate hydroxylated PA derived molecules such as 3-hydroxyphenylacetic (3-OHPA) acid and 4-hydroxyphenylacetic acid (4-OHPA). The higher activity and broader substrate specificity suggested a more active role in pathogenesis. In this work, we aimed to determine the relevance of PaaK1 activity to the killing ability of B. cenocepacia to C. elegans. Using reporter activity assays, we demonstrate that 3-OHPA activated PA degradation gene promoters of Burkholderia cenocepacia K56-2 in a paaK1-dependent manner, while 4-OHPA had no effect. We compared the pathogenicity of a paaK1 deletion mutant with that of the wild type in C. elegans and observed no differences in the killing ability of the strains. Taken together, these studies suggest that 3-OHPA, but not 4-OHPA, can induce the PA pathway and that this induction is dependent on the paaK1 gene. However, the more active PaaK1 does not play a distinct role in pathogenesis of B. cenocepacia as previously suggested.

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Defining a Structural and Kinetic Rationale for Paralogous Copies of Phenylacetate-CoA Ligases from the Cystic Fibrosis Pathogen Burkholderia cenocepacia J2315

Cited by 30 publications

References 36 publications

Insights on the regulation of the phenylacetate degradation pathway from Escherichia coli

Insights on the regulation of the phenylacetate degradation pathway from Escherichia coli

Structural Insights into Anthranilate Priming during Type II Polyketide Biosynthesis

3-Hydroxyphenylacetic Acid Induces the Burkholderia cenocepacia Phenylacetic Acid Degradation Pathway ? Toward Understanding the Contribution of Aromatic Catabolism to Pathogenesis

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