Microbial desulfonation pathways for natural and pharmacologically relevant C 3-sulfonates

Abstract

Homotaurine (3-aminopropanesulfonate), cysteate (2-amino-3-sulfopropanoate), 3-sulfolactate (2-hydroxy-3-sulfopropanoate), 2,3-dihydroxypropane-1-sulfonate (DHPS) and 3-sulfopropanoate are widespread, naturally occuring C3-sulfonates. To ournowledge, organosulfonates are degraded solely by microorganisms, which are capable of cleaving the chemically stable C-sulfonate bond. The elucidation of degradative pathways in aerobic bacteria utilizing the above mentioned C3-sulfonates as solesources of carbon and energy or, if possible, as sole sources of nitrogen, was the aim of this study.Isolates utilizing homotaurine as sole carbon and energy source or as sole nitrogen source were easily obtained. The assimilation of homotaurine-nitrogen was studied with an isolate identified as Burkholderia sp. strain N-APS2. The organism excreted3-sulfopropanoate during growth with homotaurine-nitrogen, and expressed an inducible homotaurine:2-oxoglutarate aminotransferase. The same phenomena were observed during work with the genomesequenced Cupriavidus necator H16, whichrevealed the involvement of genes and enzymes from both GABA and sulfonate metabolism: GABA permease (GabP) for homotaurine-uptake, GABA transaminase (GabT) for its deamination to 3-sulfopropanal, and succinatesemialdehydedehydrogenase (GabD1) for the oxidation of the latter to 3-sulfopropanoate, whose excretion was attributed to the sulfite/sulfonate exporter TauE.The assimilation of cysteate-nitrogen by C. necator H16 was also found to involve an initial transamination reaction. A cysteate:2-oxoglutarate aminotransferase (Coa), which might be an aspartate aminotransferase (Aoa) according to its substratepectrum, yielded 3-sulfopyruvate.Traces of the latter were found in the growth medium, together with putative 3-sulfolactate, whose formation from 3-sulfopyruvate was attributed to the activity of a putative (S)-sulfolactate dehydrogenase (SlcC). Again,TauE might be responsible for organosulfonate export; the identity of a transporter for cysteate uptake, however, is still unknown.The utilization of 3-sulfolactate as a source of carbon and energy in the genome-sequenced Roseovarius nubinhibens ISM was found to involve a largely inducible, bifurcated pathway, which allowed the desulfonation via sulfoacetaldehydecetyltransferase (Xsc) and (R)-cysteate sulfo-lyase (CuyA). A putative tripartite tricarboxylate transporter (TTT; SlcHFG) was responsible for uptake of sulfolactate, which was oxidized to 3-sulfopyruvate by a membranebound sulfolactate dehydrogenase(SlcD). 3-Sulfopyruvate, the point of bifurcation, was transaminated to cysteate in one branch, or decarboxylated to sulfoacetaldehyde in the other branch. The decarboxylating enzyme, 3-sulfopyruvate decarboxylase (ComDE) is known from thecoenzyme M biosynthetic pathway.In this study, 3-Sulfolactate was discovered as a central intermediate in the degradation of several C3-sulfonates, such as DHPS, homotaurine and 3-sulfopropanoate. In the dissimilation of racemic DHPS, three inducible DHPS dehydrogenases(HpsNOP) acted as a racemase (HpsOP) and oxidized (R)-DHPS to (R)-sulfolacate (HpsN). These enzymes were studied in R. nubinhibens ISM, which degraded the resulting sulfolactate via the bifurcated pathway described above, and in Cupriaviduspinatubonensis JMP134, which used (R)-sulfolactate sulfolyase (SuyAB) for desulfonation. Transporter candidates were available in both organisms: a putative tripartite ATP-independent periplasmic (TRAP) transporter in the marine R. nubinhibens, anda major facilitator superfamily (MFS) transporter in the terrestrial C. pinatubonensis.The utilization of homotaurine as a source of carbon and energy was studied in R. nubinhibens, whose substrate spectrum was found to include not only 3-sulfolacate and DHPS, but also homotaurine and 3-sulfopropanoate. As in the nitrogen-ssimilatory pathway, transamination and oxidation were observed, but a different transporter (HtaABCD) and aminotransferase (HtaE) became apparent. Two more novel enzymes were found for the further degradation of 3-sulfopropanoate: 3-ulfopropanoate dehydrogenase (SpuBCDA), which yielded 3-sulfopropenoate, and 3-sulfopropenoate dehydratase (SpuIJ), whose predicted product was 3-sulfolactate. Activities of Xsc and CuyA confirmed the presence of the bifurcated pathway for 3-sulfolactate degradation (see above).Work on the dissimilation of homotaurine revealed nine organisms which encoded the spu-gene cluster for 3-sulfopropanoate degradation. Rhodobacter sphaeroides 2.4.1, which grew with 3-sulfopropanoate, was presumed to degrade it via Xsc; therewere, however, no gene candidates for the sulfopyruvate decarboxylase ComDE. Instead, we found a gene encoding for a putative novel 3-sulfopyruvate decarboxylase (SpuE), which is currently under investigation.