Enhanced Production of Astaxanthin by Metabolic Engineered Isoprenoid Pathway in Escherichia coli

Abstract

The goal of this study is to increase production of astaxanthin in recombinant Escherichia coli by engineered isoprenoid pathway. We have previously reported structural and functional analysis of the astaxanthin biosynthesis genes from a marine bacterium, Paracoccus haeundaensis. The carotenoid biosynthesis gene cluster involved in astaxanthin production contained six carotenogenic genes (crtW, crtZ, crtY, crtI, crtB, and crtE genes) and recombinant E. coli harboring six carotenogenic genes from P. haeundaensis produced 400 μg/g dry cell weight (DCW) of astaxanthin. In order to increase production of astaxanthin in recombinant E. coli, we have cloned 4-hydroxy-3-methylbut-2-enyl diphosphate reductase (lytB), farnesyl diphosphate (FPP) synthase (ispA), and isopentenyl (IPP) diphossphate isomerase (idi) in the isoprenoid pathway from E. coli and coexpressed these genes in recombinant E. coli harboring the astaxanthin biosynthesis genes. This engineered E. coli strain containing both isoprenoid pathway gene and astaxanthin biosynthesis gene cluster produced 1,200 μg/g DCW of astaxanthin, resulting 3-fold increased production of astaxanthin. [14]. Even though synthetically produced astaxanthin has some drawbacks to apply commercial application, it is used for both direct and indirect food additives and coloring because naturally purified astaxanthin is very limited. In cosmetology and pharmacology, it is most demanding pigments for use as a dermal photoprotector. Astaxanthin is a pigment of high economic value and various study were carried out to increased the production of astaxanthin with the application of metabolic pathway engineering [1,18,25,28,31]. MBIC1143 [22], and Paracoccus haeundaensisIn a previous study, we isolated and characterized a marine bacterium, Paracoccus haeundaensis, which produces astaxanthin [14]. In addition, we reported the structural and functional analysis of genes encoding the astaxanthin biosynthetic enzymes; GGPP synthase (CrtE), phytoene synthase (CrtB), phytoene desaturase (CrtI), lycopene cyclase (CrtY), ß-carotene hydroxylase (CrtZ), and ß-carotene ketolase (CrtW) [15]. The individual gene of the carotenoid biosynthesis gene cluster was functionally expressed in E. coli and each gene product was purified to homogeneity. Their molecular characteristics, including enzymatic activities, were reported [16]. Furthermore, we have reported the pathways and the functions of the astaxanthin biosynthesis genes through chromatographic and spectroscopic analyses of the pigments accumulated in E. coli carrying plasmids constructed by various combinations of the carotenoid biosynthesis genes from the P. haeundaensis [17]. The astaxanthin biosynthesis pathway is summarized in Fig. 1.Enhanced production of astaxanthin in engineered microbial hosts requires optimization of the available precursors pool of isopentenyl diphosphate (IPP) and dimethylallyl diphosphate (DMAPP), and balancing the expression -Review -

Bacterial astaxanthin: production and application in aquaculture - a review

“…Nghiên cứu của Lee và Kim (2006) cho thấy, tách dòng cụm gen liên quan đến con đường tổng hợp carotenoid và tái tổ hợp vào tế bào Escherichia coli BL21(DE3) cho phép thu được 0,4 mg astaxanthin/g sinh khối khô. Lee et al, ( , 2008 không những tách dòng và biểu hiện nhóm gen tổng hợp carotenoid từ vi khuẩn Paracoccus haeundaensis, mà còn tách dòng và biểu hiện nhóm gen liên quan đến con đường tổng hợp isoprenoid. Kết quả biểu hiện trên E. coli thu được lượng astaxanthin đạt 1,2 mg/g sinh khối khô, cao gấp 3 lần so với khi chỉ tái tổ hợp nhóm gen tổng hợp carotenoid (Bảng 2).…”