Introduction of the Carotenoid Biosynthesis α-Branch Into Synechocystis sp. PCC 6803 for Lutein Production

Abstract

Lutein, made by the α-branch of the methyl-erythritol phosphate (MEP) pathway, is one of the most abundant xanthophylls in plants. It is involved in the structural stabilization of light-harvesting complexes, transfer of excitation energy to chlorophylls and photoprotection. In contrast, lutein and the α-branch of the MEP pathway are not present in cyanobacteria. In this study, we genetically engineered the cyanobacterium Synechocystis for the missing MEP α-branch resulting in lutein accumulation. A cassette comprising four Arabidopsis thaliana genes coding for two lycopene cyclases (AtLCYe and AtLCYb) and two hydroxylases (AtCYP97A and AtCYP97C) was introduced into a Synechocystis strain that lacks the endogenous, cyanobacterial lycopene cyclase cruA. The resulting synlut strain showed wild-type growth and only moderate changes in total pigment composition under mixotrophic conditions, indicating that the cruA deficiency can be complemented by Arabidopsis lycopene cyclases leaving the endogenous β-branch intact. A combination of liquid chromatography, UV-Vis detection and mass spectrometry confirmed a low but distinct synthesis of lutein at rates of 4.8 ± 1.5 nmol per liter culture at OD730 (1.03 ± 0.47 mmol mol–1 chlorophyll). In conclusion, synlut provides a suitable platform to study the α-branch of the plastidic MEP pathway and other functions related to lutein in a cyanobacterial host system.

Advances in Research on Natural Distribution, Biosynthesis, Detection, Bioactivity, and Application of Lutein

“…PCC 6803 can accumulate a large amount of lutein by changing the αbranched pathway. 23 2.6. Structure of Lutein.…”

Biochemical and Immunological implications of Lutein and Zeaxanthin

“…Cyanobacteria produce a vast range of products that mainly include β-carotene, zeaxanthin, echinenone and myxoxanthophyll [109]. In contrast to zeaxanthin, lutein and the α-branch of the MEP pathway are absent in cyanobacteria However, Lehmann et al [110] created a genetically modified cyanobacterium Synechocystis that lacks the MEP α-branch required for lutein synthesis. In this regard, a cassette encompassing four Arabidopsis thaliana genes encoding two lycopene cyclases (AtLCYe and AtLCYb) and hydroxylases (AtCYP97A and AtCYP97C) was established into a Synechocystis strain, that had a dearth of endogenous cyanobacterial lycopene cyclase cruA.…”

Main Carotenoids Produced by Microorganisms

“…GGPP molecules are in turn synthesized by the condensation of dimethylallyl pyrophosphate (DMAPP) and three molecules of isopentenyl pyrophosphate (IPP), by means of the enzyme GGPP synthase (CrtE). Mainly in eukaryotes, these precursor molecules derive from the mevalonate pathway (MEV), whilst in prokaryotes and plants they derive from the methyl-erythritol phosphate pathway (MEP), also known as the non-mevalonate pathway [34].…”

“…The production of lutein has been observed in a strain of Synechocystis sp. transformed with the previously mentioned genes [34]. In some fungi, such as X. dendrorhous, the gene crtYB encodes for a bifunctional enzyme that can perform as a phytoene synthase and as a lycopene cyclase, it combines two molecules of GGPP forming phytoene.…”

“…Finally, lutein producing bacteria has yet been found in nature, however the genetic modification of the cyanobacteria Synechocystis sp. PCC to express β-and ε-cyclase, and β-and ε-hydroxylase from Arabidopsis, results in a lutein producing strain [34].…”