Abstract: Carotenoid cleavage oxygenases (CCOs) are non-heme iron enzymes that catalyze scission of alkene groups in carotenoids and stilbenoids to form biologically important products. CCOs possess a rare four-His iron center whose resting-state structure and interaction with substrates are incompletely understood. Here, we address this knowledge gap through a comprehensive structural and spectroscopic study of three phyletically diverse CCOs. The crystal structure of a fungal stilbenoid-cleaving CCO, CAO1, reveals strong similarity between its iron center and those of carotenoid-cleaving CCOs, but with a markedly different substrate-binding cleft. These enzymes all possess a five-coordinate high-spin Fe(II) center withresting-state Fe−His bond lengths of ∼2.15 Å. This ligand set generates an iron environment more electropositive than those of other non-heme iron dioxygenases as observed by Mossbauer isomer shifts. Dioxygen (O2) does not coordinate iron in the absence of substrate. Substrates bind away (∼4.7 Å) from the iron and have little impact on its electronic structure, thus excluding coordination-triggered O2 binding. However, substrate binding does perturb the spectral properties of CCO Fe−NO derivatives, indicating proximate organic substrate and O2-binding sites, which might influence Fe−O2 interactions. Together, these data provide a robust description of the CCO iron center and its interactions with substrates and substrate mimetics that illuminates commonalities as well as subtle and profound structural differences within the CCO family.
Results from: Sui X, Weitz AC, Farquhar ER, Badiee M, Banerjee S, von Lintig J, Tochtrop GP, Palczewski K, Hendrich MP, Kiser PD. Structure and spectroscopy of alkene-cleaving dioxygenases containing an atypically coordinated non-heme iron center. Biochemistry 2017 Jun 6; 56(22):2836-2852. doi: 10.1021/acs.biochem.7b00251. Epub 2017 May 19.