Catalytic properties of the expressed acyclic carotenoid 2-ketolases from Rhodobacter capsulatus and Rubrivivax gelatinosus

Pathway Evolution by Horizontal Transfer and Positive Selection Is Accommodated by Relaxed Negative Selection upon Upstream Pathway Genes in Purple Bacterial Carotenoid Biosynthesis

“…First, positive selection at certain amino acid sites within each encoded protein might facilitate the shift from spirilloxanthin-to spheroidenone-producing pathways, the effects of which would be masked by the genewide averages analyzed here. However, the reported broad substrate specificities for upstream carotenoid biosynthetic pathway enzymes in R. gelatinosus and R. capsulatus (16,43,52,53) and the homogeneous elevation of mean d n /d s ratios (including those for the upstream-functioning gene crtB) relative to those for all other spirilloxanthinproducing bacteria by Ϸ0.15 argue against this type of selection (Table 1). Second, d n /d s ratios of carotenoid biosynthesis genes other than crtA in R. gelatinosus and H. phototrophica might be elevated due to their genetic hitchhiking with adjacent, positively selected crtA genes, whereby the selective advantages conveyed by variations in crtA outweigh the disadvantages of slightly deleterious mutations in other carotenoid biosynthetic genes.…”

“…For both organisms, lycopene production increases with increased expression of CrtI and limited production of its precursor, phytoene, whereby the lower affinity of CrtI toward neurosporene versus phytoene is counteracted by the relative lack of the latter substrate (54). In contrast, upstream pathway enzymes from both R. capsulatus and R. gelatinosus utilize a wide range of substrates from either the spheroidene or the spirilloxanthin pathway, sometimes with minimal differences in specificity (16,43,52,53). Additionally, Stickforth and Sandmann (54) observed that the substrate specificity of CrtI from R. gelatinosus could be altered by single-nucleotide mutations within the crtI gene.…”

“…strain ORS278 (17), Thiocapsa roseopersicina (30), and Rubrivivax gelatinosus (16,21,22,40,41,43,(52)(53)(54). The latter organism produces 2,2Ј-diketospirilloxanthin (lycopene derived), spheroidenone (neurosporene derived), and their precursors using the same enzymes.…”

“…Carotenoid biosynthesis has been well studied biochemically and genetically in the spheroidene-producing organisms Rhodobacter capsulatus (3,4,16,45) and Rhodobacter sphaeroides (1,31,32) and the spirilloxanthin-producing organisms Bradyrhizobium sp. strain ORS278 (17), Thiocapsa roseopersicina (30), and Rubrivivax gelatinosus (16,21,22,40,41,43,(52)(53)(54).…”

Novel Activity of Rhodobacter sphaeroides Spheroidene Monooxygenase CrtA Expressed in Escherichia coli

“…In the original host Rhodobacter strains, the monooxygenase CrtA is known to catalyze the asymmetrical introduction of one keto group at the C-2 position of spheroidene (3,4) and two keto groups at the C-2 and C-2Ј positions of spirilloxanthin (11). However, our recent studies (16,19) showed that CrtA was able to introduce one or two keto groups into acyclic nonnative substrate structures, such as -carotene and neurosporene, indicating that the substrate specificity of CrtA in Escherichia coli was more promiscuous than expected.…”