The human mitochondrial enzyme BCO2 exhibits catalytic activity toward carotenoids and apocarotenoids
Autor: | Ramkumar Srinivasagan, Nimesh Khadka, Marcin Golczak, Philip D. Kiser, Johannes von Lintig, Sepalika Bandara, Vipulkumar M. Parmar, Linda D. Thomas |
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Rok vydání: | 2020 |
Předmět: |
0301 basic medicine
Oxygenase RNA Splicing Molecular Dynamics Simulation Mitochondrion medicine.disease_cause Biochemistry Retina Dioxygenases law.invention Mice 03 medical and health sciences Zeaxanthins law Dioxygenase medicine Animals Humans Protein Isoforms Molecular Biology Escherichia coli Carotenoid chemistry.chemical_classification Binding Sites 030102 biochemistry & molecular biology Chemistry Stereoisomerism Cell Biology Metabolism Lipids Carotenoids Recombinant Proteins Mitochondria Protein Structure Tertiary 030104 developmental biology Enzyme Solubility Biocatalysis Recombinant DNA |
Zdroj: | J Biol Chem |
ISSN: | 0021-9258 |
Popis: | The enzyme β-carotene oxygenase 2 (BCO2) converts carotenoids into more polar metabolites. Studies in mammals, fish, and birds revealed that BCO2 controls carotenoid homeostasis and is involved in the pathway for vitamin A production. However, it is controversial whether BCO2 function is conserved in humans, because of a 4-amino acid long insertion caused by a splice acceptor site polymorphism. We here show that human BCO2 splice variants, BCO2a and BCO2b, are expressed as pre-proteins with mitochondrial targeting sequence (MTS). The MTS of BCO2a directed a green fluorescent reporter protein to the mitochondria when expressed in ARPE-19 cells. Removal of the MTS increased solubility of BCO2a when expressed in Escherichia coli and rendered the recombinant protein enzymatically active. The expression of the enzymatically active recombinant human BCO2a was further improved by codon optimization and its fusion with maltose-binding protein. Introduction of the 4-amino acid insertion into mouse Bco2 did not impede the chimeric enzyme's catalytic proficiency. We further showed that the chimeric BCO2 displayed broad substrate specificity and converted carotenoids into two ionones and a central C14-apocarotendial by oxidative cleavage reactions at C9,C10 and C9',C10'. Thus, our study demonstrates that human BCO2 is a catalytically competent enzyme. Consequently, information on BCO2 becomes broadly applicable in human biology with important implications for the physiology of the eyes and other tissues. |
Databáze: | OpenAIRE |
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