摘要
Carotenoids are lipophilic pigments whose light-absorbing properties and physiological functions in organisms are attributed to their conjugated double-bond system. Centric or eccentric oxidative cleavage of carotenoids is catalyzed by carotenoid cleavage dioxygenases. Humans have two CCDs that directly cleave carotenoids. These are β-carotene oxygenase 1 (BCO1) and β-carotene oxygenase 2 (BCO2).
The central region of the human retina contains concentrated levels of zeaxanthin, meso-zeaxanthin, and lutein. Collectively, these ocular carotenoids are referred to as the macula pigment. Macula pigment reduces chromatic aberration and has been shown to be photoprotective against damaging blue light. Clinical studies indicate that supplementation with these carotenoids can reduce the risk of age-related macular degeneration (AMD). Thus, there is strong advocacy for carotenoid supplementation for patients experiencing AMD. Although recommendations are based on the relationship between carotenoid intake and blood concentrations, studies in animal models indicate that this relationship is influenced by BCO2 activity and the transcription factor ISX. BCO2 converts carotenoids into more polar metabolites and ISX regulates the expression of SR-B1, which is involved in carotenoid uptake.
Aim one of this work elucidated the role of BCO2 and SR-B1 in carotenoid metabolism. We first examined BCO2’s tissue expression and determined that BCO2 is highly expressed in the small intestine of mice. Utilizing different mouse models (WT, Bco2-/- , and Isx-/- /Bco2-/- ), we demonstrated that loss of the Bco2 gene leads to increased carotenoid absorption. Additionally, increased expression of SR-B1 due to loss of Isx leads to enhanced carotenoid absorption in mice. High-performance liquid chromatography analysis showed that zeaxanthin is oxidized to ε,ε-3,3’-carotene-dione stereoisomers in the small intestine and peripheral tissues. Lastly, we generated an albino Isx-/- /Bco2-/- mouse that exhibited hypercarotenemia upon carotenoid supplementation and thus, can serve as a resourceful mammalian model for carotenoid research.
In the second aim, we address the controversy surrounding human BCO2’s enzymatic activity. We utilized recombinant isoforms of human BCO2 and mouse BCO2 to determine the biochemical factors that influence BCO2’s enzymatic activity. We demonstrated that human BCO2 isoforms BCO2a and BCO2b contain a mitochondrial targeting sequence (MTS) and that removal of the MTS results in a catalytically active human BCO2.
摘要译文
类胡萝卜素是亲脂性色素，其在生物体中的光吸收特性和生理功能归因于其共轭双键系统。类胡萝卜素的中心或偏心氧化裂解是用类胡萝卜素裂解二氧酶催化的。人类有两个直接切割类胡萝卜素的CCD。这些是β-胡萝卜素氧酶1（BCO1）和β-胡萝卜素氧酶2（BCO2）。人视网膜的中央区域含有浓缩水平的Zeaxanthin水平，中间酶 - Zexanthin和Lutein。总的来说，这些眼类胡萝卜素被称为黄斑色素。黄斑色素会减少色差，并已被证明是对破坏蓝光的光保护。临床研究表明，补充这些类胡萝卜素可以降低与年龄相关的黄斑变性（AMD）的风险。因此，对于患有AMD的患者，对类胡萝卜素补充有很大的倡导。尽管建议基于类胡萝卜素摄入与血液浓度之间的关系，但动物模型的研究表明，这种关系受BCO2活性和转录因子ISX的影响。BCO2将类胡萝卜素转化为更多的极性代谢产物，ISX调节SR-B1的表达，SR-B1参与类胡萝卜素摄取。IAM这项工作阐明了BCO2和SR-B1在类胡萝卜素代谢中的作用。我们首先检查了BCO2的组织表达，并确定BCO2在小鼠的小肠中高度表达。利用不同的鼠标模型（wt，bco2 - / - 和isx - / - /bco2 - / - ），我们证明了失去的损失BCO2基因导致类胡萝卜素吸收增加。另外，由于ISX的损失导致SR-B1的表达增加导致小鼠的类胡萝卜素吸收增强。高性能的液相色谱分析表明，在小肠和外围组织中，Zeaxanthin被氧化为ε，ε-3,3,-胡萝卜素二酮立体异构体。最后，我们产生了一个白化病ISX - / - /bco2 - / - 小鼠，在补充类胡萝卜素时表现出高氧基血症，因此可以用作用于类胡萝卜素研究的哺乳动物模型。在第二个目标中，我们解决了围绕人BCO2酶活性的争议。我们利用人BCO2和小鼠BCO2的重组同工型来确定影响BCO2酶促活性的生化因子。我们证明了人BCO2同工型BCO2A和BCO2B包含线粒体靶向序列（MTS），并且去除MTS会导致催化活性的人类BCO2。
Thomas, Linda D.. Characterization of Carotenoid Metabolism and Its Scavenger Enzyme, BCO2[D]. US: Case Western Reserve University, 2023