Unknown

Dataset Information

0

Structural and Functional Characterization of the PaaI Thioesterase from Streptococcus pneumoniae Reveals a Dual Specificity for Phenylacetyl-CoA and Medium-chain Fatty Acyl-CoAs and a Novel CoA-induced Fit Mechanism.


ABSTRACT: PaaI thioesterases are members of the TE13 thioesterase family that catalyze the hydrolysis of thioester bonds between coenzyme A and phenylacetyl-CoA. In this study we characterize the PaaI thioesterase from Streptococcus pneumoniae (SpPaaI), including structural analysis based on crystal diffraction data to 1.8-Å resolution, to reveal two double hotdog domains arranged in a back to back configuration. Consistent with the crystallography data, both size exclusion chromatography and small angle x-ray scattering data support a tetrameric arrangement of thioesterase domains in solution. Assessment of SpPaaI activity against a range of acyl-CoA substrates showed activity for both phenylacetyl-CoA and medium-chain fatty-acyl CoA substrates. Mutagenesis of putative active site residues reveals Asn(37), Asp(52), and Thr(68) are important for catalysis, and size exclusion chromatography analysis and x-ray crystallography confirm that these mutants retain the same tertiary and quaternary structures, establishing that the reduced activity is not a result of structural perturbations. Interestingly, the structure of SpPaaI in the presence of CoA provides a structural basis for the observed substrate specificity, accommodating a 10-carbon fatty acid chain, and a large conformational change of up to 38 Å in the N terminus, and a loop region involving Tyr(38)-Tyr(39). This is the first time PaaI thioesterases have displayed a dual specificity for medium-chain acyl-CoAs substrates and phenylacetyl-CoA substrates, and we provide a structural basis for this specificity, highlighting a novel induced fit mechanism that is likely to be conserved within members of this enzyme family.

SUBMITTER: Khandokar YB 

PROVIDER: S-EPMC4722464 | biostudies-literature | 2016 Jan

REPOSITORIES: biostudies-literature

altmetric image

Publications

Structural and Functional Characterization of the PaaI Thioesterase from Streptococcus pneumoniae Reveals a Dual Specificity for Phenylacetyl-CoA and Medium-chain Fatty Acyl-CoAs and a Novel CoA-induced Fit Mechanism.

Khandokar Yogesh B YB   Srivastava Parul P   Sarker Subir S   Swarbrick Crystall M D CMD   Aragao David D   Cowieson Nathan N   Forwood Jade K JK  

The Journal of biological chemistry 20151104 4


PaaI thioesterases are members of the TE13 thioesterase family that catalyze the hydrolysis of thioester bonds between coenzyme A and phenylacetyl-CoA. In this study we characterize the PaaI thioesterase from Streptococcus pneumoniae (SpPaaI), including structural analysis based on crystal diffraction data to 1.8-Å resolution, to reveal two double hotdog domains arranged in a back to back configuration. Consistent with the crystallography data, both size exclusion chromatography and small angle  ...[more]

Similar Datasets

| S-EPMC3086008 | biostudies-literature
| S-EPMC3195577 | biostudies-literature
| S-EPMC3336778 | biostudies-literature
| S-EPMC5733585 | biostudies-literature
| S-EPMC4242439 | biostudies-literature
| S-EPMC8401288 | biostudies-literature
| S-EPMC3525216 | biostudies-literature
| S-EPMC1189074 | biostudies-literature
| S-EPMC9836253 | biostudies-literature
| S-EPMC5587920 | biostudies-literature