Elsevier

Gene

Volume 189, Issue 2, 21 April 1997, Pages 169-174
Gene

Cloning, sequencing and expressing the carotenoid biosynthesis genes, lycopene cyclase and phytoene desaturase, from the aerobic photosynthetic bacterium Erythrobacter longus sp. strain Och101 in Escherichia coli

https://doi.org/10.1016/S0378-1119(96)00788-3Get rights and content

Abstract

Two genes which encode the enzymes lycopene cyclase and phytoene desaturase in the aerobic photosynthetic bacterium Erythrobacter longus sp. strain Och101 have been cloned and sequenced. The gene for lycopene cyclase, designated crtY, was expressed in a strain of Escherichia coli which contained the crtE, B, I and Z genes encoding geranylgeranyl pyrophosphate synthase, phytoene synthase, phytoene desaturase, and β-carotene hydroxylase, respectively. As a result, zeaxanthin production was observed in E. coli transformants. In addition, expression of the E. longus gene crtI for phytoene desaturase in E. coli containing crtE and B resulted in the accumulation of lycopene in transformants. Zeaxanthin and lycopene were also determined by mass spectrum. Nucleotide sequence similarities between E. longus crtY gene and other microbial lycopene cyclase genes are 40.2% (Erwinia herbicola), 37.4% (Erwinia uredovora) and 22.9% (Synechococcus sp.), and those between phytoene desaturase genes are 50.3% (E. herbicola), 54.7% (E. uredovora) and 39.6% (Rhodobacter capsulatus).

Introduction

Carotenoids are an important group of natural pigments which are widely distributed in living organisms. They can function as protective compounds against photooxidative damage, as light harvesting pigments and as pigments which impact color to the living tissues. Certain cyclic carotenoids, such as β-carotene, are precursors of vitamin A in animals and are of current interest as nutritional factors important for cancer prevention (Lambert et al., 1990).

Genes which encode the biosynthetic enzymes for carotenoids have been cloned and sequenced from the photosynthetic bacterium Rhodobacter capsulatus (Armstrong et al., 1989), and the phytopathogenic bacteria Erwinia herbicola and Erwinia uredovora (Hundle et al., 1993; Misawa et al., 1990). However, genes which encode lycopene cyclase (Fig. 1), a key enzyme which converts the acyclic carotenoid lycopene into the cyclic carotenoid β-carotene, have not been isolated from photosynthetic bacteria. In the oxygenic cyanobacteria, where β-carotene is an essential component of the photosystem, a lycopene cyclase gene has recently been characterized (Cunningham et al., 1993, Cunningham et al., 1994).

Here we describe the cloning and sequencing of genes encoding phytoene desaturase and lycopene cyclase from the aerobic photosynthetic bacterium Erythrobacter longus in an effort to understand the organization and expression of the carotenoid biosynthesis genes in this organism. E. longus has an interesting carotenoid composition and has been found to produce about 20 different kinds of carotenoids such as β-carotene and monocyclic carotenoids such as rubixanthan (Takaichi et al., 1990).

Section snippets

Cloning of the lycopene cyclase and phytoene desaturase genes

A gene library was constructed from chromosomal Erythrobacter longus DNA which was partially digested with Sau3AI and size fractionated to give fragments which were 6–20 kb in size. The DNA was ligated into the BamHI site of the vector λEMBL3. The resulting gene library contained about 1.5×105 (95%) individual clones. To clone the phytoene desaturase genes from E. longus, a 1 kb BglII-PstI fragment containing the phytoene desaturase gene from Erwinia herbicola was used as a probe for

Conclusions

  • 1.

    Two carotenoid biosynthetic genes, crtY and crtI, encoding lycopene cyclase and phytoene desaturase respectively have been cloned from the aerobic photosynthetic bacterium Erythrobacter longus.

  • 2.

    The nucleotide sequence similarities of E. longus crtY gene are 40.2%, 37.4% and 22.9% to the corresponding genes in E. herbicola, E. uredovora and Synechococcus sp., respectively.

  • 3.

    The nucleotide sequence similarities of E. longus crtI gene is 50.3%, 54.7% and 39.6% to the corresponding genes in E.

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There are more references available in the full text version of this article.

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