Dataset Information

X-ray crystal structure of a malonate-semialdehyde dehydrogenase from Pseudomonas sp. strain AAC.

ABSTRACT: The NAD-dependent malonate-semialdehyde dehydrogenase KES23460 from Pseudomonas sp. strain AAC makes up half of a bicistronic operon responsible for ?-alanine catabolism to produce acetyl-CoA. The KES23460 protein has been heterologously expressed, purified and used to generate crystals suitable for X-ray diffraction studies. The crystals belonged to space group P212121 and diffracted X-rays to beyond 3?Å resolution using the microfocus beamline of the Australian Synchrotron. The structure was solved using molecular replacement, with a monomer from PDB entry 4zz7 as the search model.

SUBMITTER: Wilding M

PROVIDER: S-EPMC5287376 | biostudies-literature | 2017 Jan

REPOSITORIES: biostudies-literature

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X-ray crystal structure of a malonate-semialdehyde dehydrogenase from Pseudomonas sp. strain AAC.

Wilding Matthew M Scott Colin C Peat Thomas S TS Newman Janet J

Acta crystallographica. Section F, Structural biology communications 20170101 Pt 1

The NAD-dependent malonate-semialdehyde dehydrogenase KES23460 from Pseudomonas sp. strain AAC makes up half of a bicistronic operon responsible for β-alanine catabolism to produce acetyl-CoA. The KES23460 protein has been heterologously expressed, purified and used to generate crystals suitable for X-ray diffraction studies. The crystals belonged to space group P212121 and diffracted X-rays to beyond 3 Å resolution using the microfocus beamline of the Australian ...[more]

PMID: 28045390

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Project description:We previously isolated the transaminase KES23458 from Pseudomonas sp. strain AAC as a promising biocatalyst for the production of 12-aminododecanoic acid, a constituent building block of nylon-12. Here, we report the subsequent characterization of this transaminase. It exhibits activity with a broad substrate range which includes ?-, ?-, and ?-amino acids, as well as ?,?-diamines and a number of other industrially relevant compounds. It is therefore a prospective candidate for the biosynthesis of a range of polyamide monomers. The crystal structure of KES23458 revealed that the protein forms a dimer containing a large active site pocket and unusual phosphorylated histidine residues. To infer the physiological role of the transaminase, we expressed, purified, and characterized a dehydrogenase from the same operon, KES23460. Unlike the transaminase, the dehydrogenase was shown to be quite selective, catalyzing the oxidation of malonic acid semialdehyde, formed from ?-alanine transamination via KES23458. In keeping with previous reports, the dehydrogenase was shown to catalyze both a coenzyme A (CoA)-dependent reaction to form acetyl-CoA and a significantly slower CoA-independent reaction to form acetate. These findings support the original functional assignment of KES23458 as a ?-alanine transaminase. However, a seemingly well-adapted active site and promiscuity toward unnatural compounds, such as 12-aminododecanoic acid, suggest that this enzyme could perform multiple functions for Pseudomonas sp. strain AAC.We describe the characterization of an industrially relevant transaminase able to metabolize 12-aminododecanoic acid, a constituent building block of the widely used polymer nylon-12, and we report the biochemical and structural characterization of the transaminase protein. A physiological role for this highly promiscuous enzyme is proposed based on the characterization of a related gene from the host organism. Molecular dynamics simulations were carried out to compare the conformational changes in the transaminase protein to better understand the determinants of specificity in the protein. This study makes a substantial contribution that is of interest to the broad biotechnology and enzymology communities, providing insights into the catalytic activity of an industrially relevant biocatalyst as well as the biological function of this operon.

Dataset Information

X-ray crystal structure of a malonate-semialdehyde dehydrogenase from Pseudomonas sp. strain AAC.

Publications

X-ray crystal structure of a malonate-semialdehyde dehydrogenase from Pseudomonas sp. strain AAC.

Similar Datasets

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