4-Sulfomuconolactone Hydrolases from Hydrogenophaga intermedia S1 and Agrobacterium radiobacter S2

Abstract

The 4-carboxymethylen-4-sulfo-but-2-en-olide (4-sulfomuconolactone) hydrolases from Hydrogenophaga intermedia strain S1 and Agrobacterium radiobacter strain S2 are part of a modified protocatechuate pathway responsible for the degradation of 4-sulfocatechol. In both strains, the hydrolase-encoding genes occur downstream of those encoding the enzymes that catalyze the lactonization of 3-sulfomuconate. The deduced amino acid sequences of the 4-sulfomuconolactone hydrolases demonstrated the highest degree of sequence identity to 2-pyrone-4,6-dicarboxylate hydrolases, which take part in the meta cleavage pathway of protocatechuate. The 4-sulfomuconolactone hydrolases did not convert 2-pyrone-4,6-dicarboxylate, and the 2-pyrone-4,6-dicarboxylate hydrolase from Sphingomonas paucimobilis SYK-6 did not convert 4-sulfomuconolactone. Nevertheless, the presence of highly conserved histidine residues in the 4-sulfomuconolactone and the 2-pyrone-4,6-dicarboxylate hydrolases and some further sequence similarities suggested that both enzymes belong to the metallo-dependent hydrolases (the "amidohydrolase superfamily"). The 4-sulfomuconolactone hydrolases were heterologously expressed as His-tagged enzyme variants. Gel filtration experiments suggested that the enzymes are present as monomers in solution, with molecular weights of approximately 33,000 to 35,000. 4-Sulfomuconolactone was converted by sulfomuconolactone hydrolases to stoichiometric amounts of maleylacetate and sulfite. The 4-sulfomuconolactone hydrolases from both strains showed pH optima at pH 7 to 7.5 and rather similar catalytic constant (k cat /K M )values. The suggested 4-sulfocatechol pathway from 4-sulfocatechol to maleylacetate was confirmed by in situ nuclear magnetic resonance analysis using the recombinantly expressed enzymes.

Two Enzymes of a Complete Degradation Pathway for Linear Alkylbenzenesulfonate (LAS) Surfactants: 4-Sulfoacetophenone Baeyer-Villiger Monooxygenase and 4-Sulfophenylacetate Esterase in Comamonas testosteroni KF-1

“…Second, how is 4-sulfophenol converted to (desulfonated) central metabolites in strain KF-1? We found evidence for the involvement of 4-sulfocatechol as the substrate for a desulfonative ortho-ring cleavage pathway (40,42,45), as has been characterized in other bacteria (9,15,16); however, we have preliminary evidence to suggest that the pathway may also (or instead?) involve 2-hydroxyquinol (1,2, 4-trihydroxybenzene) ortho-ring cleavage.…”

“…1) (for details and structures, see reference 42). The lower 3-C 4 -SPC pathway, i.e., after the proposed BVMO-esterase reaction sequence and from 4-sulfophenol further via aromatic ring cleavage to central metabolites, is thought to proceed via 4-sulfophenol 2-monooxygenation to 4-sulfocatechol, followed by 4-sulfocatechol orthoring cleavage (9,15,16) (see schematic representation in Fig. 1) (for details and structures, see reference 42).…”

Cloning and functional analysis of the genes coding for 4-aminobenzenesulfonate 3,4-dioxygenase from Hydrogenophaga sp. PBC

“…The subsequent identification of its additional components provided a very crucial piece of information to elucidate the 4-ABS degradation pathway. Combining the data from several 4-ABS-biodegradation studies (Contzen et al, 2001;Feigel & Knackmuss, 1993;Gan et al, 2011a;Halak et al, 2006Halak et al, , 2007Hammer et al, 1996) and the data from this study, a complete pathway for 4-ABS biodegradation is proposed (Fig. 5).…”

“…Instead of liberating the sulfonate group before ring cleavage to form catechol, the sulfonate group is retained and 4-sulfocatechol is oxidized to generate a new compound, 3-sulfomuconate. The genes and enzymes involved in the conversion of 4-sulfocatechol to maleylacetate by Hydrogenophaga intermedia S1 have been characterized in great detail via heterologous expression in recombinant Escherichia coli (Halak et al, 2006(Halak et al, , 2007Hammer et al, 1996).…”

Genome Sequence of Hydrogenophaga sp. Strain PBC, a 4-Aminobenzenesulfonate-Degrading Bacterium

“…PBC strains were auxotrophic for biotin and p-aminobenzoic acid, which may explain the roles of their respective helper strains in the biodegradation of 4-ABS. To date, the molecular mechanism of 4-ABS degradation has been studied in great detail in this genus via an approach involving transposon mutagenesis and enzymology (2,4,(7)(8)(9). The recent identification of the genetic components for the oxidation of 4-ABS to 4-sulfocatechol from Hydrogenophaga sp.…”