The complete genome sequence of Roseobacter denitrificans reveals a mixotrophic rather than photosynthetic metabolism

Abstract

Purple aerobic anoxygenic phototrophs (AAPs) are the only organisms known to capture light energy to enhance growth only in the presence of oxygen but do not produce oxygen. The highly adaptive AAPs compose more than 10% of the microbial community in some euphotic upper ocean waters and are potentially major contributors to the fixation of the greenhouse gas CO2. We present the complete genomic sequence and feature analysis of the AAP Roseobacter denitrificans, which reveal clues to its physiology. The genome lacks genes that code for known photosynthetic carbon fixation pathways, and most notably missing are genes for the Calvin cycle enzymes ribulose bisphosphate carboxylase (RuBisCO) and phosphoribulokinase. Phylogenetic evidence implies that this absence could be due to a gene loss from a RuBisCO-containing alpha-proteobacterial ancestor. We describe the potential importance of mixotrophic rather than autotrophic CO2 fixation pathways in these organisms and suggest that these pathways function to fix CO2 for the formation of cellular components but do not permit autotrophic growth. While some genes that code for the redox-dependent regulation of photosynthetic machinery are present, many light sensors and transcriptional regulatory motifs found in purple photosynthetic bacteria are absent.

FIG. 1.

Circular representation of the Roseobacter denitrificans OCh 114 chromosome (maps of plasmids pTB1, pTB2, pTB3, and pTB4 are found in Fig. S1 in the supplemental material). The different rings represent (from outer to inner) all genes and insertion elements, which are color coded by functional category (rings 1 and 2), deviation from average G+C content (ring 3), and GC skew (ring 4). Color codes are as follows: turquoise indicates small-molecule biosynthesis, yellow indicates central or intermediary metabolism, black indicates energy metabolism, red indicates signal transduction, lavender indicates DNA metabolism, blue indicates transcription, purple indicates protein synthesis/fate, dark green indicates surface-associated features, gray indicates miscellaneous features, pink indicates phage and insertion elements, light green indicates unknown function, orange indicates hypothetical proteins, and brown indicates pseudogenes.

FIG. 2.

16S ribosomal tree for the α-proteobacterial lineage using genomes available in the NCBI database. The tree was constructed using maximum likelihood methods. RuBisCO-containing taxa are indicated in red text, with putative RuBisCO-containing nodes highlighted with red dots. Both clades of aerobic phototrophic bacteria (AAP) are highlighted within green boxes. This tree illustrated the widespread character of RuBisCO among α-proteobacteria as well as its conspicuous absence within the AAPs. Note that without the vast oversampling among AAP lineages, a higher percentage of sequenced α-proteobacteria contain RuBisCO than do not.

FIG. 3.

Proposed mixotrophic CO₂ assimilation pathway in Roseobacter denitrificans. CO₂ is taken up from the environment or supplied by the action of CO dehydrogenase. Carbonic anhydrase converts CO₂ to HCO₃⁻, which is then used by PEP carboxylase and pyruvate carboxylase. OAA is either entered into the TCA cycle or converted to aspartate, which can be converted to several amino acids and pyrimidine nucleotides. Although the assimilated carbon can be lost by the action of 2-ketoglutarate dehydrogenase, it will be maintained in TCA cycle intermediates that are recruited into other pathways.

FIG. 4.

Photosynthesis gene cluster arrangement in the purple bacteria Rubrivivax gelatinosus, Roseobacter denitrificans, and Rhodobacter capsulatus. Genes are presented as arrows indicating their direction of transcription. Genes are colored as listed: chlorophyll biosynthesis (bch) in green, carotenoid biosynthesis (crt) in orange, reaction centers and light-harvesting complexes (puf and puh) in purple, regulatory proteins in blue, and uncharacterized genes in white. All other gene colors are unique for clarity. Lines illustrate gene rearrangement, and arrows illustrate the reversal of large superoperonal clusters.