A genomic perspective on the potential of Actinobacillus succinogenes for industrial succinate production

Abstract

Background: Succinate is produced petrochemically from maleic anhydride to satisfy a small specialty chemical market. If succinate could be produced fermentatively at a price competitive with that of maleic anhydride, though, it could replace maleic anhydride as the precursor of many bulk chemicals, transforming a multi-billion dollar petrochemical market into one based on renewable resources. Actinobacillus succinogenes naturally converts sugars and CO2 into high concentrations of succinic acid as part of a mixed-acid fermentation. Efforts are ongoing to maximize carbon flux to succinate to achieve an industrial process.

Results: Described here is the 2.3 Mb A. succinogenes genome sequence with emphasis on A. succinogenes's potential for genetic engineering, its metabolic attributes and capabilities, and its lack of pathogenicity. The genome sequence contains 1,690 DNA uptake signal sequence repeats and a nearly complete set of natural competence proteins, suggesting that A. succinogenes is capable of natural transformation. A. succinogenes lacks a complete tricarboxylic acid cycle as well as a glyoxylate pathway, and it appears to be able to transport and degrade about twenty different carbohydrates. The genomes of A. succinogenes and its closest known relative, Mannheimia succiniciproducens, were compared for the presence of known Pasteurellaceae virulence factors. Both species appear to lack the virulence traits of toxin production, sialic acid and choline incorporation into lipopolysaccharide, and utilization of hemoglobin and transferrin as iron sources. Perspectives are also given on the conservation of A. succinogenes genomic features in other sequenced Pasteurellaceae.

Conclusions: Both A. succinogenes and M. succiniciproducens genome sequences lack many of the virulence genes used by their pathogenic Pasteurellaceae relatives. The lack of pathogenicity of these two succinogens is an exciting prospect, because comparisons with pathogenic Pasteurellaceae could lead to a better understanding of Pasteurellaceae virulence. The fact that the A. succinogenes genome encodes uptake and degradation pathways for a variety of carbohydrates reflects the variety of carbohydrate substrates available in the rumen, A. succinogenes's natural habitat. It also suggests that many different carbon sources can be used as feedstock for succinate production by A. succinogenes.

Figure 1

Nucleotide frequency in A. succinogenes USS repeats.

Figure 2

A. succinogenes central metabolic network based on annotation of metabolic genes. Four-digit numbers are Asuc_ORF (locus tags) numbers and are followed by E.C. numbers. Hyphenated locus tag numbers indicate that the enzyme is encoded by several successive genes. Reaction names: see additional file 2: Table S3. Arrow and number colors: black, product function assumed; green, putative function assumed; blue, probable function assumed; red, possible function assumed. Metabolites: AcCoA, acetyl-CoA; Ace, acetate; AcP, acetyl-phosphate; Ald, acetaldehyde; Cit, citrate; EtOH, ethanol; E4P, erythrose-4-phosphate; For, formate; Fum, fumarate; F1,6P, fructose-1,6-bisphosphate; F6P, fructose-6-phosphate; Glc, glucose; G1,3P, glycerate-1,3-bisphosphate; G3P, glyceraldehyde-3-phosphate; G6P, glucose-6-phosphate; Icit, Isocitrate; αKG, α-ketoglutarate; Mal, malate; OAA, oxaloacetate; PEP, phosphoenolpyruvate; 2PG, 2-phosphoglycerate; 3PG, 3-phosphoglycerate; Pyr, pyruvate; Q⁺, menaquinone; QOH, menaquinol; R5P, ribose-5-phosphate, Ru5P, ribulose-5-phosphate, Suc, succinate; SucCoA, succinyl-CoA; S7P, sedoheptulose-7-phosphate; X5P, xylulose-5-phosphate.

Figure 3

A. succinogenes uptake and degradation pathways for sugars other than glucose. See additional file 2: Table S1 for more details. Arrow and number colors: black, product function assumed; green, putative function assumed; blue, probable function assumed; red, possible function assumed. Bold arrows, glycolytic and pentose phosphate pathways as described in Figure 2 and additional file 2: Table S3; Outer-membrane triangle, maltoporin. Numbers refer to enzymes or transporters described in additional file 2: Table S1. Metabolites: 5K4DG, 5-dehydro-4-deoxy-D-glucarate; 5 KG, 5-ketogluconate; Ara, L-arabinose; A6P, ascorbate-6-phosphate; DHAP, dihydroxyacetone phosphate; F1P, fructose-1-phosphate; Frc, fructose; Gal, galactose; Gal1P, galactose-1-phosphate; Galte, galactarate; Glcte, glucarate; Gly, glycerol; Gte, gluconate; Gt6P, gluconate-6-phosphate; G1P, glucose-1-phosphate; βG6P, β-glucoside-6-phosphate; Gt6P, gluconate-6-phosphate; Ido, idonate; KDPG, 2-keto-3-deoxy-6-phosphogluconate; Lac, lactose; Mal, maltose; Man6P, mannose-6-phosphate; MOH1P, mannitol-1-phosphate; Pec, pectin; Rib, ribose; Ribu, ribulose; S6P, sucrose-6-phosphate; SOH6P, sorbitol-1-phosphate; Xyl, xylose; Xylu, xylulose. Other abbreviations are as in Figure 2.