Search

Deinococcus radiodurans strain R1 utilizes multiple antioxidants including a unique carotenoid, deinoxanthin, to fight again oxidative stress. Most of the enzymes involved in the deinoxanthin biosynthetic pathway have been identified. However, the enzyme catalysing the synthesis of geranylgeranyl diphosphate ( GGPP), which is a precursor of carotenoid biosynthesis, has yet to be identified. Two putative isoprenyl diphosphate synthases ( IPPS) homologues ( DR1395 and DR932) were screened out by analysis of conserved amino acid regions, and their biochemical functions were investigated. Gene mutation, gene expression in Escherichia coli and analysis of carotenoid products were used to investigate the functions of these candidates. The results suggested that DR1395 encodes the protein for GGPP synthesis. Site-directed mutant analysis indicated that the amino acid composition of and around the first aspartate-rich motif is vital for GGPP synthase function. Significance and Impact of the Study Deinococcus radiodurans strain R1 produces a unique carotenoid product, deinoxanthin, as an antioxidant. In this study, DR1395 was identified as the gene encoding geranylgeranyl diphosphate synthase ( GGPPS) for entrance to deinoxanthin biosynthesis in D. radiodurans. Moreover, site-directed mutagenesis studies on DR1395 identified the effect of amino acid composition of the aspartate-rich motif on the production of this carotenoid. This study demonstrated the entrance step in the deinoxanthin biosynthetic pathway. These results can be useful in genetic engineering strategies for deinoxanthin production including enhancement of GGPPS gene expression in D. radiodurans. [ABSTRACT FROM AUTHOR]

Aims: To evaluate the antioxidant effect of carotenoids from Deinococcus radiodurans on protein. Methods and Results: Deinococcus radiodurans strain R1 (ATCC 13939) and its mutant strain R1ΔcrtB were used for this study. The total carotenoids (R1ex) from D. radiodurans were obtained by extraction with acetone/methanol (7 : 2, by vol), and their antioxidant activity was measured using the DPPH˙ (2,2-diphenyl-1-picrylhydrazyl) system. The protein oxidation level, in vitro and in the cell, was measured using the DNPH (2,4-dinitrophenyl hydrazine) method. The carotenoid extract R1ex scavenged 40·2% DPPH˙ radicals compared to β-carotene (31·7%) at a concentration of 0·5 mg ml−1. The intracellular level of protein oxidation in mutant R1ΔcrtB, which does not contain carotenoid, was 0·0212 mmol mg−1 protein which is significantly greater than that in the wild type (0·0169 mmol mg−1 protein) following the treatment with H2O2. The purified major carotenoid product (deinoxanthin) from the wild type showed a greater inhibition of oxidative damage in bovine serum albumin than lycopene or lutein. Conclusions: Carotenoids prevent protein oxidation and contribute to the resistance to cell damage in D. radiodurans. Significance and Impact of the Study: Our results provide the evidence that carotenoids can protect proteins in D. radiodurans against oxidative stress. [ABSTRACT FROM AUTHOR]