Global genome analysis of the shikimic acid pathway reveals greater gene loss in host-associated than in free-living bacteria

Abstract

Background: A central tenet in biochemistry for over 50 years has held that microorganisms, plants and, more recently, certain apicomplexan parasites synthesize essential aromatic compounds via elaboration of a complete shikimic acid pathway, whereas metazoans lacking this pathway require a dietary source of these compounds. The large number of sequenced bacterial and archaean genomes now available for comparative genomic analyses allows the fundamentals of this contention to be tested in prokaryotes. Using Hidden Markov Model profiles (HMM profiles) to identify all known enzymes of the pathway, we report the presence of genes encoding shikimate pathway enzymes in the hypothetical proteomes constructed from the genomes of 488 sequenced prokaryotes.

Results: Amongst free-living prokaryotes most Bacteria possess, as expected, genes encoding a complete shikimic acid pathway, whereas of the culturable Archaea, only one was found to have a complete complement of recognisable enzymes in its predicted proteome. It may be that in the Archaea, the primary amino-acid sequences of enzymes of the pathway are highly divergent and so are not detected by HMM profiles. Alternatively, structurally unrelated (non-orthologous) proteins might be performing the same biochemical functions as those encoding recognized genes of the shikimate pathway. Most surprisingly, 30% of host-associated (mutualistic, commensal and pathogenic) bacteria likewise do not possess a complete shikimic acid pathway. Many of these microbes show some degree of genome reduction, suggesting that these host-associated bacteria might sequester essential aromatic compounds from a parasitised host, as a 'shared metabolic adaptation' in mutualistic symbiosis, or obtain them from other consorts having the complete biosynthetic pathway. The HMM results gave 84% agreement when compared against data in the highly curated BioCyc reference database of genomes and metabolic pathways.

Conclusions: These results challenge the conventional belief that the shikimic acid pathway is universal and essential in prokaryotes. The possibilities that non-orthologous enzymes catalyse reactions in this pathway (especially in the Archaea), or that there exist specific uptake mechanisms for the acquisition of shikimate intermediates or essential pathway products, warrant further examination to better understand the precise metabolic attributes of host-beneficial and pathogenic bacteria.

Figure 1

Schematic representation of the shikimic acid pathway. The first step is an aldol condensation between phosphoenol pyruvate [1] and erythrose-4-phosphate [2] catalyzed by 3-deoxy-D-arabinoheptulosonate-7-phosphate synthase (DAHP, or aroG/F/H according to E. coli nomenclature, EC 4.1.2.15) to synthesize 3-deoxy-D-arabino-heptulosonate-7-phosphate [3]. The second step is carried out by 3-dehydroquinate synthase (DHQ synthase, aroB, EC 4.6.1.3), affording 3-dehydroquinate [4]. The third step synthesizes 3-dehydroshikimate [5] catalyzed by 3-dehydroquinate dehydratase (DH-quinase, aroD, EC 4.2.1.10). Step 4 is catalyzed by shikimate dehydrogenase (Shikimate_DH, aroE, EC 1.1.1.25) to synthesize shikimate [6]. Steps 5 and 6 are catalyzed by shikimate kinase (SKI, aroK, EC 2.7.1.71) and 5-enolpyruvylshikimate-3-phosphate synthase (EPSP, aroA, EC 2.5.1.19) to give shikimate-3-phosphate [7] and 5-enolpyruvylshikimate-3-phosphate [8], respectively. The last reaction, catalyzed by chorismate synthase (aroC, EC 4.6.1.4) affords the final product of the pathway, chorismate [9].

Figure 2

HMM analysis of the shikimic acid pathway in the predicted proteomes of 147 'free-living' Bacteria shows that most have the complete pathway and only a few are missing one or more enzymes in the pathway.

Figure 3

HMM analysis showing the distribution of enzymes forming the shikimic acid pathway in the predicted proteomes of 147 'free-living' Bacteria; there is no pattern to which enzymes are missing.

Figure 4

HMM analysis showing the distribution of enzymes forming the shikimic acid pathway in the predicted proteomes of 250 host-associated Bacteria that can be cultured in the laboratory ex hospite shows that there is a nearly equal loss of enzymes across the pathway.

Figure 5

HMM analysis of the distribution of enzymes forming the shikimic acid pathway in the predicted proteomes of 46 host-associated Bacteria that cannot be cultured ex hospite shows that there is a nearly equal loss of enzymes across the pathway.

Figure 6

HMM analysis of the shikimic acid pathway in the predicted proteomes of 46 Archaea shows that nearly all of these prokaryotes are missing only one or two enzymes in the pathway.

Figure 7

HMM analysis showing the distribution of enzymes forming the shikimic acid pathway in the predicted proteomes of 46 Archaea; nearly all of these prokaryotes are missing either the first enzyme in the pathway, DAHP synthase, or the fifth enzyme, shikimate kinase.