Characterization of the Role of β-Carotene 9,10-Dioxygenase in Macular Pigment Metabolism

Darwin Babino,Grzegorz Palczewski,M Airanthi K Widjaja-Adhi,Philip D Kiser,Marcin Golczak,Johannes Von Lintig

doi:10.1074/jbc.m115.668822

Darwin Babino, Grzegorz Palczewski + Show 4 more

Open Access

https://doi.org/10.1074/jbc.m115.668822

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Abstract

A family of enzymes collectively referred to as carotenoid cleavage oxygenases is responsible for oxidative conversion of carotenoids into apocarotenoids, including retinoids (vitamin A and its derivatives). A member of this family, the β-carotene 9,10-dioxygenase (BCO2), converts xanthophylls to rosafluene and ionones. Animals deficient in BCO2 highlight the critical role of the enzyme in carotenoid clearance as accumulation of these compounds occur in tissues. Inactivation of the enzyme by a four-amino acid-long insertion has recently been proposed to underlie xanthophyll concentration in the macula of the primate retina. Here, we focused on comparing the properties of primate and murine BCO2s. We demonstrate that the enzymes display a conserved structural fold and subcellular localization. Low temperature expression and detergent choice significantly affected binding and turnover rates of the recombinant enzymes with various xanthophyll substrates, including the unique macula pigment meso-zeaxanthin. Mice with genetically disrupted carotenoid cleavage oxygenases displayed adipose tissue rather than eye-specific accumulation of supplemented carotenoids. Studies in a human hepatic cell line revealed that BCO2 is expressed as an oxidative stress-induced gene. Our studies provide evidence that the enzymatic function of BCO2 is conserved in primates and link regulation of BCO2 gene expression with oxidative stress that can be caused by excessive carotenoid supplementation.

Highlights

BCO2 converts xanthophylls in rodents, but it is controversial whether this role is conserved in primates
Structural Comparison between Rodent and Primate BCO2s—Besides BCO2, the human genome encodes two other carotenoid cleavage oxygenase (CCO) family members, retinal pigment epithelium protein of 65 kDa (RPE65) and BCO1, both of which are critical for human retinoid metabolism [24]
Sequence alignments revealed that the most pronounced difference between human BCO2 and other family members is a long N-terminal leader sequence for mitochondrial import [13] (Fig. 1). This N-terminal extension is present in other primate BCO2s, e.g. the predicted macaque BCO2 (MaBCO2), but is absent in murine BCO2 (MuBCO2) [11, 13]

Summary

Background

BCO2 converts xanthophylls in rodents, but it is controversial whether this role is conserved in primates. Results: Recombinant primate BCO2 displays enzymatic activity and is expressed as an oxidative stress-induced mitochondrial protein. Our studies provide evidence that the enzymatic function of BCO2 is conserved in primates and link regulation of BCO2 gene expression with oxidative stress that can be caused by excessive carotenoid supplementation. Genetic studies in animals suggest that the ␤-carotene 9,10dioxygenase (BCO2) plays a critical role in controlling carotenoid tissue levels and preventing excess accumulation of these compounds. We utilized human cell lines and mouse models, respectively, to study the transcriptional regulation and function of this protein in carotenoid homeostasis of blood and tissues. The picture that emerges verifies the critical role that BCO2s play in this process and indicates that oxidative stress in chronic disease induces BCO2 and carotenoid breakdown in tissues and blood

Experimental Procedures

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