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Molecular basis for the substrate specificity of quorum signal synthases.

ABSTRACT: In several Proteobacteria, LuxI-type enzymes catalyze the biosynthesis of acyl-homoserine lactones (AHL) signals using S-adenosyl-l-methionine and either cellular acyl carrier protein (ACP)-coupled fatty acids or CoA-aryl/acyl moieties as progenitors. Little is known about the molecular mechanism of signal biosynthesis, the basis for substrate specificity, or the rationale for donor specificity for any LuxI member. Here, we present several cocrystal structures of BjaI, a CoA-dependent LuxI homolog that represent views of enzyme complexes that exist along the reaction coordinate of signal synthesis. Complementary biophysical, structure-function, and kinetic analysis define the features that facilitate the unusual acyl conjugation with S-adenosylmethionine (SAM). We also identify the determinant that establishes specificity for the acyl donor and identify residues that are critical for acyl/aryl specificity. These results highlight how a prevalent scaffold has evolved to catalyze quorum signal synthesis and provide a framework for the design of small-molecule antagonists of quorum signaling.

SUBMITTER: Dong SH

PROVIDER: S-EPMC5576808 | biostudies-literature | 2017 Aug

REPOSITORIES: biostudies-literature

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Molecular basis for the substrate specificity of quorum signal synthases.

Dong Shi-Hui SH Frane Nicole D ND Christensen Quin H QH Greenberg E Peter EP Nagarajan Rajesh R Nair Satish K SK

Proceedings of the National Academy of Sciences of the United States of America 20170807 34

In several <i>Proteobacteria</i>, LuxI-type enzymes catalyze the biosynthesis of acyl-homoserine lactones (AHL) signals using <i>S</i>-adenosyl-l-methionine and either cellular acyl carrier protein (ACP)-coupled fatty acids or CoA-aryl/acyl moieties as progenitors. Little is known about the molecular mechanism of signal biosynthesis, the basis for substrate specificity, or the rationale for donor specificity for any LuxI member. Here, we present several cocrystal structures of BjaI, a CoA-depend ...[more]

PMID: 28784791

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Project description:Oxalate decarboxylase (OxDC) catalyzes the conversion of oxalate into formate and carbon dioxide in a remarkable reaction that requires manganese and dioxygen. Previous studies have shown that replacing an active-site loop segment Ser(161)-Glu(162)-Asn(163)-Ser(164) in the N-terminal domain of OxDC with the cognate residues Asp(161)-Ala(162)-Ser-(163)-Asn(164) of an evolutionarily related, Mn-dependent oxalate oxidase gives a chimeric variant (DASN) that exhibits significantly increased oxidase activity. The mechanistic basis for this change in activity has now been investigated using membrane inlet mass spectrometry (MIMS) and isotope effect (IE) measurements. Quantitative analysis of the reaction stoichiometry as a function of oxalate concentration, as determined by MIMS, suggests that the increased oxidase activity of the DASN OxDC variant is associated with only a small fraction of the enzyme molecules in solution. In addition, IE measurements show that C-C bond cleavage in the DASN OxDC variant proceeds via the same mechanism as in the wild-type enzyme, even though the Glu(162) side chain is absent. Thus, replacement of the loop residues does not modulate the chemistry of the enzyme-bound Mn(II) ion. Taken together, these results raise the possibility that the observed oxidase activity of the DASN OxDC variant arises from an increased level of access of the solvent to the active site during catalysis, implying that the functional role of Glu(162) is to control loop conformation. A 2.6 Å resolution X-ray crystal structure of a complex between oxalate and the Co(II)-substituted ΔE162 OxDC variant, in which Glu(162) has been deleted from the active site loop, reveals the likely mode by which the substrate coordinates the catalytically active Mn ion prior to C-C bond cleavage. The "end-on" conformation of oxalate observed in the structure is consistent with the previously published V/K IE data and provides an empty coordination site for the dioxygen ligand that is thought to mediate the formation of Mn(III) for catalysis upon substrate binding.

Dataset Information

Molecular basis for the substrate specificity of quorum signal synthases.

Publications

Molecular basis for the substrate specificity of quorum signal synthases.

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