7v28
Crystal Structure of phthalate dioxygenase in complex with terephthalate
Structural highlights
Publication Abstract from PubMedPhthalate, a plasticizer, endocrine disruptor, and potential carcinogen, is degraded by a variety of bacteria. This degradation is initiated by phthalate dioxygenase (PDO), a Rieske oxygenase (RO) that catalyzes the dihydroxylation of phthalate to a dihydrodiol. PDO has long served as a model for understanding ROs despite a lack of structural data. Here we purified PDOKF1 from Comamonas testosteroni KF1 and found that it had an apparent kcat/Km for phthalate of 0.58 +/- 0.09 muM(-1)s(-1), over 25-fold greater than for terephthalate. The crystal structure of the enzyme at 2.1 A resolution revealed that it is a hexamer comprising two stacked alpha3 trimers, a configuration not previously observed in RO crystal structures. We show that within each trimer, the protomers adopt a head-to-tail configuration typical of ROs. The stacking of the trimers is stabilized by two extended helices, which make the catalytic domain of PDOKF1 larger than that of other characterized ROs. Complexes of PDOKF1 with phthalate and terephthalate revealed that Arg207 and Arg244, two residues on one face of the active site, position these substrates for regiospecific hydroxylation. Consistent with their roles as determinants of substrate specificity, substitution of either residue with alanine yielded variants that did not detectably turnover phthalate. Together, these results provide critical insights into a pollutant-degrading enzyme that has served as a paradigm for ROs and facilitate the engineering of this enzyme for bioremediation and biocatalytic applications. Molecular insights into substrate recognition and catalysis by phthalate dioxygenase from Comamonas testosteroni.,Mahto JK, Neetu N, Waghmode B, Kuatsjah E, Sharma M, Sircar D, Sharma AK, Tomar S, Eltis LD, Kumar P J Biol Chem. 2021 Nov 17;297(6):101416. doi: 10.1016/j.jbc.2021.101416. PMID:34800435[1] From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine. References
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