Dataset Information

Substrate Specificity of Purified Recombinant Chicken ?-Carotene 9',10'-Oxygenase (BCO2).

ABSTRACT: Provitamin A carotenoids are oxidatively cleaved by ?-carotene 15,15'-dioxygenase (BCO1) at the central 15-15' double bond to form retinal (vitamin A aldehyde). Another carotenoid oxygenase, ?-carotene 9',10'-oxygenase (BCO2) catalyzes the oxidative cleavage of carotenoids at the 9'-10' bond to yield an ionone and an apo-10'-carotenoid. Previously published substrate specificity studies of BCO2 were conducted using crude lysates from bacteria or insect cells expressing recombinant BCO2. Our attempts to obtain active recombinant human BCO2 expressed in Escherichia coli were unsuccessful. We have expressed recombinant chicken BCO2 in the strain E. coli BL21-Gold (DE3) and purified the enzyme by cobalt ion affinity chromatography. Like BCO1, purified recombinant chicken BCO2 catalyzes the oxidative cleavage of the provitamin A carotenoids ?-carotene, ?-carotene, and ?-cryptoxanthin. Its catalytic activity with ?-carotene as substrate is at least 10-fold lower than that of BCO1. In further contrast to BCO1, purified recombinant chicken BCO2 also catalyzes the oxidative cleavage of 9-cis-?-carotene and the non-provitamin A carotenoids zeaxanthin and lutein, and is inactive with all-trans-lycopene and ?-apocarotenoids. Apo-10'-carotenoids were detected as enzymatic products by HPLC, and the identities were confirmed by LC-MS. Small amounts of 3-hydroxy-?-apo-8'-carotenal were also consistently detected in BCO2-?-cryptoxanthin reaction mixtures. With the exception of this activity with ?-cryptoxanthin, BCO2 cleaves specifically at the 9'-10' bond to produce apo-10'-carotenoids. BCO2 has been shown to function in preventing the excessive accumulation of carotenoids, and its broad substrate specificity is consistent with this.

SUBMITTER: Dela Sena C

PROVIDER: S-EPMC4938182 | biostudies-literature | 2016 Jul

REPOSITORIES: biostudies-literature

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Substrate Specificity of Purified Recombinant Chicken β-Carotene 9',10'-Oxygenase (BCO2).

Dela Seña Carlo C Sun Jian J Narayanasamy Sureshbabu S Riedl Kenneth M KM Yuan Yan Y Curley Robert W RW Schwartz Steven J SJ Harrison Earl H EH

The Journal of biological chemistry 20160503 28

Provitamin A carotenoids are oxidatively cleaved by β-carotene 15,15'-dioxygenase (BCO1) at the central 15-15' double bond to form retinal (vitamin A aldehyde). Another carotenoid oxygenase, β-carotene 9',10'-oxygenase (BCO2) catalyzes the oxidative cleavage of carotenoids at the 9'-10' bond to yield an ionone and an apo-10'-carotenoid. Previously published substrate specificity studies of BCO2 were conducted using crude lysates from bacteria or insect cells expressing recombinant BCO2. Our atte ...[more]

PMID: 27143479

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Project description:BackgroundPlasma cholesterol is one of the strongest risk factors associated with the development of atherosclerotic cardiovascular disease (ASCVD) and myocardial infarction. Human studies suggest that elevated plasma β-carotene is associated with reductions in circulating cholesterol and the risk of myocardial infarction. The molecular mechanisms underlying these observations are unknown.ObjectiveThe objective of this study was to determine the impact of dietary β-carotene and the activity of β-carotene oxygenase 1 (BCO1), which is the enzyme responsible for the conversion of β-carotene to vitamin A, on circulating cholesterol concentration.MethodsIn our preclinical study, we compared the effects of a 10-d intervention with a diet containing 50 mg/kg of β-carotene on plasma cholesterol in 5-wk-old male and female C57 Black 6 wild-type and congenic BCO1-deficient mice. In our clinical study, we aimed to determine whether 5 common small nucleotide polymorphisms located in the BCO1 locus affected serum cholesterol concentrations in a population of young Mexican adults from the Universities of San Luis Potosí and Illinois: A Multidisciplinary Investigation on Genetics, Obesity, and Social-Environment (UP AMIGOS) cohort.ResultsUpon β-carotene feeding, Bco1-/- mice accumulated >20-fold greater plasma β-carotene and had ∼30 mg/dL increased circulating total cholesterol (P < 0.01) and non-HDL cholesterol (P < 0.01) than wild-type congenic mice. Our results in the UP AMIGOS cohort show that the rs6564851 allele of BCO1, which has been linked to BCO1 enzymatic activity, was associated with a reduction in 10 mg/dL total cholesterol concentrations (P = 0.009) when adjusted for vitamin A and carotenoid intakes. Non-HDL-cholesterol concentration was also reduced by 10 mg/dL when the data were adjusted for vitamin A and total carotenoid intakes (P = 0.002), or vitamin A and β-carotene intakes (P = 0.002).ConclusionsOverall, our results in mice and young adults show that BCO1 activity impacts circulating cholesterol concentration, linking vitamin A formation with the risk of developing ASCVD.

Dataset Information

Substrate Specificity of Purified Recombinant Chicken ?-Carotene 9',10'-Oxygenase (BCO2).

Publications

Substrate Specificity of Purified Recombinant Chicken β-Carotene 9',10'-Oxygenase (BCO2).

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