Characterization of a Gene Cluster Involved in 4-Chlorocatechol Degradation byPseudomonas reinekeiMT1

Abstract

Pseudomonas reinekei MT1 has previously been reported to degrade 4-and 5-chlorosalicylate by a pathway with 4-chlorocatechol, 3-chloromuconate, 4-chloromuconolactone, and maleylacetate as intermediates, and a gene cluster channeling various salicylates into an intradiol cleavage route has been reported. We now report that during growth on 5-chlorosalicylate, besides a novel (chloro)catechol 1,2-dioxygenase, C12O ccaA , a novel (chloro)muconate cycloisomerase, MCI ccaB , which showed features not yet reported, was induced. This cycloisomerase, which was practically inactive with muconate, evolved for the turnover of 3-substituted muconates and transforms 3-chloromuconate into equal amounts of cis-dienelactone and protoanemonin, suggesting that it is a functional intermediate between chloromuconate cycloisomerases and muconate cycloisomerases. The corresponding genes, ccaA (C12O ccaA ) and ccaB (MCI ccaB ), were located in a 5.1-kb genomic region clustered with genes encoding trans-dienelactone hydrolase (ccaC) and maleylacetate reductase (ccaD) and a putative regulatory gene, ccaR, homologous to regulators of the IclR-type family. Thus, this region includes genes sufficient to enable MT1 to transform 4-chlorocatechol to 3-oxoadipate. Phylogenetic analysis showed that C12O ccaA and MCI ccaB are only distantly related to previously described catechol 1,2-dioxygenases and muconate cycloisomerases. Kinetic analysis indicated that MCI ccaB and the previously identified C12O salD , rather than C12O ccaA , are crucial for 5-chlorosalicylate degradation. Thus, MT1 uses enzymes encoded by a completely novel gene cluster for degradation of chlorosalicylates, which, together with a gene cluster encoding enzymes for channeling salicylates into the ortho-cleavage pathway, form an effective pathway for 4-and 5-chlorosalicylate mineralization.

Degradation of paracetamol by pure bacterial cultures and their microbial consortium

“…4 (continued) carbon, nitrogen, and energy. Members of the Pseudomonas are environmental organisms known for their ability to degrade aromatic compounds of environmental concern (Cámara et al 2009;Neumann et al 2004). However, except for P. aeruginosa (Gusseme et al 2011), the genus has not included any known paracetamol degraders until now.…”

Bacterial degradation of chlorophenols and their derivatives

“…Other genes present on this cluster include cph C-I , cphC -II, cphF -I , cphF -II, Cph B, Cph X, CphR and Cph S. The genes cphC -I and cph C-II encode putative monooxygenase, whereas cph F-1 and cph F-11 encode putative maleylacetate reductase and cph B encodes a NADH:flavin adenine dinucleotide oxidoreductase. The roles of the remaining genes in the cph gene cluster have yet to be determined [ 112 ] .…”

Novel metal-binding site of Pseudomonas reinekei MT1 trans-dienelactone hydrolase

“…One example is Pseudomonas reinekei MT1, which degrades 4-chlorocatechol via a route, which harbors a novel enzyme termed trans-dienelactone hydrolase (trans-DLH) instead of a typical DLH (Fig. 1) [5,6].…”