Oxalyl-Coenzyme A Reduction to Glyoxylate Is the Preferred Route of Oxalate Assimilation in Methylobacterium extorquens AM1

Abstract

Oxalate catabolism is conducted by phylogenetically diverse organisms, including Methylobacterium extorquens AM1. Here, we investigate the central metabolism of this alphaproteobacterium during growth on oxalate by using proteomics, mutant characterization, and 13 C-labeling experiments. Our results confirm that energy conservation proceeds as previously described for M. extorquens AM1 and other characterized oxalotrophic bacteria via oxalyl-coenzyme A (oxalyl-CoA) decarboxylase and formyl-CoA transferase and subsequent oxidation to carbon dioxide via formate dehydrogenase. However, in contrast to other oxalate-degrading organisms, the assimilation of this carbon compound in M. extorquens AM1 occurs via the operation of a variant of the serine cycle as follows: oxalyl-CoA reduction to glyoxylate and conversion to glycine and its condensation with methylene-tetrahydrofolate derived from formate, resulting in the formation of C3 units. The recently discovered ethylmalonyl-CoA pathway operates during growth on oxalate but is nevertheless dispensable, indicating that oxalyl-CoA reductase is sufficient to provide the glyoxylate required for biosynthesis. Analysis of an oxalyl-CoA synthetase- and oxalyl-CoA-reductase-deficient double mutant revealed an alternative, although less efficient, strategy for oxalate assimilation via one-carbon intermediates. The alternative process consists of formate assimilation via the tetrahydrofolate pathway to fuel the serine cycle, and the ethylmalonyl-CoA pathway is used for glyoxylate regeneration. Our results support the notion that M. extorquens AM1 has a plastic central metabolism featuring multiple assimilation routes for C1 and C2 substrates, which may contribute to the rapid adaptation of this organism to new substrates and the eventual coconsumption of substrates under environmental conditions.

Systems-level Proteomics of Two Ubiquitous Leaf Commensals Reveals Complementary Adaptive Traits for Phyllosphere Colonization

“…Exported formate can subsequently be oxidized by the periplasmic dehydrogenase Fdh3, which was up-regulated during plant colonization. Previous characterization of the oxalate metabolism in M. extorquens AM1 revealed that reduction of oxalyl-CoA to glyoxylate is the preferred route of oxalate assimilation and that NAD(P) ϩ transhydrogenase for redox balance was highly expressed in cells grown on oxalate (52). In the present study, NAD(P) ϩ transhydrogenase was induced upon plant colonization in M. extorquens PA1, however, the corresponding proteins for assimilatory reduction to glyoxylate were down-regulated, indicating that oxalate oxidation is rather used for energy conservation than for carbon assimilation (supplemental Table S3).…”

A Link between Arabinose Utilization and Oxalotrophy in Bradyrhizobium japonicum

“…Several intermediates of the L-arabinose degradation pathway (L-2-keto-3-deoxyarabonate, glycolaldehyde, and glyoxylate) as well as oxalate could be identified by a metabolomics approach (using nanoflow ion-pair RP-HPLC coupled with nanospray high-resolution MS with a split-free nano-LC Ultra system connected to an LTQ-Orbitrap mass spectrometer). Sampling, quenching, and central metabolite extraction were carried out as described by Schneider and colleagues (32). The data were normalized to the biomass produced in the cultures.…”

The Family Methylobacteriaceae

“…The oxalate-grown organisms also contained L-serine-glyoxylate aminotransferase and hydroxypyruvate reductase, indicating that synthesis of C 3 -compounds from oxalate occurred by a variant of the serine pathway ''used by Pseudomonas AM1 during growth on C 1 -compounds'' (Blackmore and Quayle 1970). This metabolism was subsequently confirmed for other Methylobacterium species and for Mtb extorquens strain AM1 (Schneider et al 2012), and the mechanism of oxalate oxidation and assimilation is available on the MetaCyc database for Mtb extorquens (http:// metacyc.org/META/NEW-IMAGE?type=NIL&object=PWY-6696) and Mtb chloromethanicum (http://biocyc.org/MCHL440085/ NEW-IMAGE?object=Oxalate-Degradation).…”