Identification of a fourth family of lycopene cyclases in photosynthetic bacteria - PubMed (original) (raw)

Identification of a fourth family of lycopene cyclases in photosynthetic bacteria

Julia A Maresca et al. Proc Natl Acad Sci U S A. 2007.

Abstract

A fourth and large family of lycopene cyclases was identified in photosynthetic prokaryotes. The first member of this family, encoded by the cruA gene of the green sulfur bacterium Chlorobium tepidum, was identified in a complementation assay with a lycopene-producing strain of Escherichia coli. Orthologs of cruA are found in all available green sulfur bacterial genomes and in all cyanobacterial genomes that lack genes encoding CrtL- or CrtY-type lycopene cyclases. The cyanobacterium Synechococcus sp. PCC 7002 has two homologs of CruA, denoted CruA and CruP, and both were shown to have lycopene cyclase activity. Although all characterized lycopene cyclases in plants are CrtL-type proteins, genes orthologous to cruP also occur in plant genomes. The CruA- and CruP-type carotenoid cyclases are members of the FixC dehydrogenase superfamily and are distantly related to CrtL- and CrtY-type lycopene cyclases. Identification of these cyclases fills a major gap in the carotenoid biosynthetic pathways of green sulfur bacteria and cyanobacteria.

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Conflict of interest statement

The authors declare no conflict of interest.

Figures

Fig. 1.

Fig. 1.

Pigments produced in E. coli lycopene complementation strains. The absorption spectra of numbered peaks are shown on the right. Pigments extracted from E. coli BL21 (DE3) cells harboring plasmids pAC-LYC and pET16b, the empty vector (A); plasmids pAC-LYC and pCTLY, which encodes C. tepidum cruA (B); and plasmids pAC-LYC and p3-SLP, which encodes Synechococcus sp. PCC 7002 cruP (C). Peak 1 is lycopene; peak 2 is γ-carotene; and peak 3 is β-carotene.

Fig. 2.

Fig. 2.

Neighbor-joining phylogenetic tree of four classes of lycopene cyclases. Bold lines indicate genes or gene products that have been either genetically or biochemically characterized.

Fig. 3.

Fig. 3.

Insertional inactivation of cruA and cruP. Constructs for insertional inactivation of cruA in C. tepidum (A) cruA in Synechococcus sp. PCC 7002 (B) cruP in Synechococcus sp. PCC 7002 (C). All three constructs produced homozygous mutants, as shown by PCR across the insertions. The locus amplified is indicated above the figure. (D) Lanes 1, wild-type C. tepidum; lanes 2 and 3, two isolates of the C. tepidum cruA::aadA mutant. (E) Lanes 1, wild-type Synechococcus sp. PCC 7002; lanes 2, Synechococcus cruA::aacC1 mutant. (F) Lanes 1, wild-type Synechococcus sp. PCC 7002; lanes 2, Synechococcus cruP::aphA mutant. M, DNA size markers. The numbers at the left are sizes in kilobases.

Fig. 4.

Fig. 4.

HPLC elution profiles, monitored at 491 nm, of carotenoids extracted from wild-type (black lines) and cruA mutant strains (gray lines). (A) Carotenoids extracted from C. tepidum strains and normalized to BChl c content. Peak 1, OH-chlorobactene glucoside laurate; peak 2, chlorobactene; peak 3, lycopene; peak 4, 1′,2′-dihydrochlorobactene; peak 5, γ-carotene. (B) Carotenoids from Synechococcus sp. PCC 7002 strains, normalized to Chl a content. Peak 3, lycopene; peak 5, γ-carotene; peak 6, β-carotene; peak 7, cryptoxanthin; peak 8, Chl a. ∗, unidentified carotenoids with lycopene chromophores.

Fig. 5.

Fig. 5.

Reaction(s) catalyzed by CruA and CruP. CruA and CruP from Synechococcus sp. PCC 7002 have both activities. However, the overall activity of CruP is lower in Synechococcus sp. PCC 7002, and CruP may also have less activity in converting γ-carotene to β-carotene.

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