Novel Pathway of Salicylate Degradation by Streptomyces sp. Strain WA46

Abstract

A novel salicylate-degrading Streptomyces sp., strain WA46, was identified by UV fluorescence on solid minimal medium containing salicylate; trace amounts of gentisate were detected by high-pressure liquid chromatography when strain WA46 was grown with salicylate. PCR amplification of WA46 DNA with degenerate primers for gentisate 1,2-dioxygenase (GDO) genes produced an amplicon of the expected size. Sequential PCR with nested GDO primers was then used to identify a salicylate degradation gene cluster in a plasmid library of WA46 chromosomal DNA. The nucleotide sequence of a 13.5-kb insert in recombinant plasmid pWD1 (which was sufficient for the complete degradation of salicylate) showed that nine putative open reading frames (ORFs) (sdgABCDEFGHR) were involved. Plasmid pWD1 derivatives disrupted in each putative gene were transformed into Streptomyces lividans TK64. Disruption of either sdgA or sdgC blocked salicylate degradation; constructs lacking sdgD accumulated gentisate. Cell extracts from Escherichia coli DH5␣ transformants harboring pUC19 that expressed each of the sdg ORFs showed that conversions of salicylate to salicylylcoenzyme A (CoA) and salicylyl-CoA to gentisyl-CoA required SdgA and SdgC, respectively. SdgA required CoA and ATP as cofactors, while NADH was required for SdgC activity; SdgC was identified as salicylyl-CoA 5-hydroxylase. Gentisyl-CoA underwent spontaneous cleavage to gentisate and CoA. SdgA behaved as a salicylyl-CoA ligase despite showing amino acid sequence similarity to an AMP-ligase. SdgD was identified as a GDO. These results suggest that Streptomyces sp. strain WA46 degrades salicylate by a novel pathway via a CoA derivative. Two-dimensional polyacrylamide gel electrophoresis and reverse transcriptase-PCR studies indicated that salicylate induced expression of the sdg cluster.

Cloning and Characterization of the Biosynthetic Gene Cluster for Tomaymycin, an SJG-136 Monomeric Analog

“…TomO shares sequence similarity (25% identity) with salicylyl-coenzyme A (CoA) 5-hydroxylase shown to catalyze the hydroxylation of salicylyl-CoA to gentisyl-CoA (15). This reaction is identical to the 5-hydroxylation of anthranilic acid with the exception that the carboxylate of anthranilic acid is esterified by CoA (15). Therefore, TomO is proposed to catalyze the 5-hydroxylation of anthranilic acid in the tomaymycin gene cluster.…”

“…Hydroxylation and O methylation of anthranilic acid are likely catalyzed by the remaining ORFs identified in the tomaymycin gene clusters tomE, tomF, tomG, and tomO. TomO shares sequence similarity (25% identity) with salicylyl-coenzyme A (CoA) 5-hydroxylase shown to catalyze the hydroxylation of salicylyl-CoA to gentisyl-CoA (15). This reaction is identical to the 5-hydroxylation of anthranilic acid with the exception that the carboxylate of anthranilic acid is esterified by CoA (15).…”

Microbial expression profiles in the rhizosphere of willows depend on soil contamination

“…Salicylate is derived from the amino acid phenylalanine and is degraded through the catechol or gentisate pathways (Ishiyama et al, 2004). Salicylate-degrading bacteria were previously found to be enriched beneath willows in comparison with other vegetation (Schmidt et al, 2000).…”

Metabolism of 2-hydroxy-1-naphthoic acid and naphthalene via gentisic acid by distinctly different sets of enzymes in Burkholderia sp. strain BC1

“…Salicylic acid has been studied most widely and was reported to be metabolized by the action of either salicylate 1-hydroxylase via catechol (Pinyakong et al, 2003;Simon et al, 1993), salicylate 5-hydroxylase via gentisic acid (Zhou et al, 2001) or direct ortho ring-cleavage dioxygenase via 2-oxohepta-3,5-dienedioic acid (Hintner et al, 2001). Salicylic acid was also metabolized through gentisic acid via salicylylCoA (Ishiyama et al, 2004) and through catechol via the decarboxylase-mediated transformation product phenol (Iwasaki et al, 2010). By contrast, hydroxynaphthoic acids were metabolized by the action of hydroxynaphthoic acid hydroxylase via dihydroxynaphthalene (Pinyakong et al, 2000), by direct ortho ring-cleavage of 1H2NA via trans-29 -carboxybenzalpyruvate (Iwabuchi & Harayama, 1997) or by direct meta ring-cleavage of 1H2NA and 2H1NA via 2,2-dicarboxychromene and 2-hydroxychromene-2-glyoxylic acid, respectively (Mallick et al, 2007;Roy et al, 2012).…”