7WZD

Crystal Structure of cis-4,5-dihydrodiol phthalate dehydrogenase from Comamonas testosteroni KF1

7WZD の概要

• エントリーDOI: 10.2210/pdb7wzd/pdb
• 分子名称: 4,5-dihydroxyphthalate dehydrogenase, GLYCEROL (3 entities in total)
• 機能のキーワード: dehydrogenase, bacterial protein, nad cofactor, oxidoreductase
• 由来する生物種: Comamonas testosteroni KF-1
• タンパク質・核酸の鎖数: 2
• 化学式量合計: 86369.63

構造登録者

Sharma, M.,Mahto, J.K.,Kumar, P. (登録日: 2022-02-17, 公開日: 2022-09-14, 最終更新日: 2023-11-29)

主引用文献

Mahto, J.K.,Sharma, M.,Neetu, N.,Kayastha, A.,Aggarwal, S.,Kumar, P.
Conformational flexibility enables catalysis of phthalate cis-4,5-dihydrodiol dehydrogenase.
Arch.Biochem.Biophys., 727:109314-109314, 2022

PubMed Abstract: Phthalate cis-4,5-dihydrodiol dehydrogenase (PhtC), the second enzyme of the phthalate catabolic pathway, catalyzes the dehydrogenation of cis-4,5-dihydrodiol phthalate (DDP). Here, we report the structural and biochemical characterization of PhtC from Comamonas testosteroni KF1 (PhtC). With biochemical experiments, we have determined the enzyme's catalytic efficiency (k/K) with DDP as 2.6 ± 0.5 Ms, over 10-fold higher than with cis-3,4-dihydrodiol phthalate (CDP). To understand the structural basis of these reactions, the crystal structures of PhtC in apo-form, the binary complex with NAD, and the ternary complex with NAD and 3-hydroxybenzoate (3HB) were determined. These crystal structures reveal that the binding of 3HB induces a conformational change in the substrate-binding loop. This conformational change causes the opening of the NAD  binding site while trapping the 3HB. The PhtC crystal structures show that the catalytic domain of PhtC is larger than that of other structurally characterized homologs and does not align with other cis-diol dehydrogenases. Structural and mutational analysis of the substrate-binding loop residues, Arg164 and Glu167 establish that conformational flexibility of this loop is necessary for positioning the substrate in a catalytically competent pose, as substitution of either of these residues to Ala did not yield the dehydrogenation activity. Further, based on the crystal structures of PhtC and related structural homologs, a reaction mechanism is proposed. Finally, with the biochemical analysis of a variant M251LPhtC, the broader substrate specificity of this enzyme is explained.

PubMed: 35667443
DOI: 10.1016/j.abb.2022.109314

実験手法

X-RAY DIFFRACTION (2.8 Å)