Evolution of Enzymes Required for Biodegradation of Pentachlorophenol in Sphingobium Chlorophenolicum

2007 ◽  
Author(s):  
Shelley D. Copley
Keyword(s):  
Sphingobium Chlorophenolicum

scholarly journals Maintenance Role of a Glutathionyl-Hydroquinone Lyase (PcpF) in Pentachlorophenol Degradation by Sphingobium chlorophenolicum ATCC 39723

2008 ◽  
Vol 190 (23) ◽  
pp. 7595-7600 ◽  
Author(s):  
Yan Huang ◽  
Randy Xun ◽  
Guanjun Chen ◽  
Luying Xun
Keyword(s):  
Degradation Rate ◽  
Tricarboxylic Acid Cycle ◽  
Degradation Pathway ◽  
Tricarboxylic Acid ◽  
Metabolic Intermediate ◽  
Reductive Dehalogenase ◽  
Cell Extracts ◽  
Toxic Pollutant ◽  
Sphingobium Chlorophenolicum

ABSTRACT Pentachlorophenol (PCP) is a toxic pollutant. Its biodegradation has been extensively studied in Sphingobium chlorophenolicum ATCC 39723. All enzymes required to convert PCP to a common metabolic intermediate before entering the tricarboxylic acid cycle have been characterized. One of the enzymes is tetrachloro-p-hydroquinone (TeCH) reductive dehalogenase (PcpC), which is a glutathione (GSH) S-transferase (GST). PcpC catalyzes the GSH-dependent conversion of TeCH to trichloro-p-hydroquinone (TriCH) and then to dichloro-p-hydroquinone (DiCH) in the PCP degradation pathway. PcpC is susceptible to oxidative damage, and the damaged PcpC produces glutathionyl (GS) conjugates, GS-TriCH and GS-DiCH, which cannot be further metabolized by PcpC. The fate and effect of GS-hydroquinone conjugates were unknown. A putative GST gene (pcpF) is located next to pcpC on the bacterial chromosome. The pcpF gene was cloned, and the recombinant PcpF was purified. The purified PcpF was able to convert GS-TriCH and GS-DiCH conjugates to TriCH and DiCH, respectively. The GS-hydroquinone lyase reactions catalyzed by PcpF are rather unusual for a GST. The disruption of pcpF in S. chlorophenolicum made the mutant lose the GS-hydroquinone lyase activities in the cell extracts. The mutant became more sensitive to PCP toxicity and had a significantly decreased PCP degradation rate, likely due to the accumulation of the GS-hydroquinone conjugates inside the cell. Thus, PcpF played a maintenance role in PCP degradation and converted the GS-hydroquinone conjugates back to the intermediates of the PCP degradation pathway.

scholarly journals Structures of the Inducer-Binding Domain of Pentachlorophenol-Degrading Gene Regulator PcpR from Sphingobium chlorophenolicum

2014 ◽  
Vol 15 (11) ◽  
pp. 20736-20752 ◽  
Author(s):  
Robert Hayes ◽  
Timothy Moural ◽  
Kevin Lewis ◽  
David Onofrei ◽  
Luying Xun ◽  
...  
Keyword(s):  
Binding Domain ◽  
Sphingobium Chlorophenolicum

Rhizoremediation of pentachlorophenol by Sphingobium chlorophenolicum ATCC 39723

Chemosphere ◽  
2007 ◽  
Vol 68 (5) ◽  
pp. 864-870 ◽  
Author(s):  
R.I. Dams ◽  
G.I. Paton ◽  
K. Killham
Keyword(s):  
Sphingobium Chlorophenolicum

scholarly journals Residues His172 and Lys238 are Essential for the Catalytic Activity of the Maleylacetate Reductase from Sphingobium chlorophenolicum Strain L-1

2017 ◽  
Vol 7 (1) ◽  
Author(s):  
Lifeng Chen ◽  
Ed S. Krol ◽  
Meena K. Sakharkar ◽  
Haseeb A. Khan ◽  
Abdullah S. Alhomida ◽  
...  
Keyword(s):  
Catalytic Activity ◽  
Sphingobium Chlorophenolicum ◽  
Maleylacetate Reductase

scholarly journals Organization and Regulation of Pentachlorophenol-Degrading Genes in Sphingobium chlorophenolicum ATCC 39723

2002 ◽  
Vol 184 (17) ◽  
pp. 4672-4680 ◽  
Author(s):  
Mian Cai ◽  
Luying Xun
Keyword(s):  
Transcriptional Regulator ◽  
Degradation Pathway ◽  
Gene Products ◽  
Regulator Gene ◽  
New Genes ◽  
Reductase Gene ◽  
Sphingomonas Chlorophenolica ◽  
Sphingobium Chlorophenolicum ◽  
Maleylacetate Reductase ◽  
Regulator Genes

ABSTRACT The first three enzymes of the pentachlorophenol (PCP) degradation pathway in Sphingobium chlorophenolicum (formerly Sphingomonas chlorophenolica) ATCC 39723 have been characterized, and the corresponding genes, pcpA, pcpB, and pcpC, have been individually cloned and sequenced. To search for new genes involved in PCP degradation and map the physical locations of the pcp genes, a 24-kb fragment containing pcpA and pcpC was completely sequenced. A putative LysR-type transcriptional regulator gene, pcpM, and a maleylacetate reductase gene, pcpE, were identified upstream of pcpA. pcpE was found to play a role in PCP degradation. pcpB was not found on the 24-kb fragment. The four gene products PcpB, PcpC, PcpA, and PcpE were responsible for the metabolism of PCP to 3-oxoadipate in ATCC 39723, and inactivational mutation of each gene disrupted the degradation pathway. The organization of the pcp genes is unusual because the four PCP-degrading genes, pcpA, pcpB, pcpC, and pcpE, were found to be located at four discrete locations. Two hypothetical LysR-type regulator genes, pcpM and pcpR, have been identified; pcpM was not required, but pcpR was essential for the induction of pcpB, pcpA, and pcpE. The coinducers of PcpR were PCP and other polychlorinated phenols. The expression of pcpC was constitutive. Thus, the organization and regulation of the genes involved in PCP degradation to 3-oxoadipate were documented.

scholarly journals Genome Shuffling Improves Degradation of the Anthropogenic Pesticide Pentachlorophenol by Sphingobium chlorophenolicum ATCC 39723

2004 ◽  
Vol 70 (4) ◽  
pp. 2391-2397 ◽  
Author(s):  
MingHua Dai ◽  
Shelley D. Copley
Keyword(s):  
Type Strain ◽  
Wild Type Strain ◽  
Constitutive Expression ◽  
Strain Analysis ◽  
Genome Shuffling ◽  
Original Strain ◽  
Negative Bacterium ◽  
Wild Type ◽  
Enhanced Resistance ◽  
Sphingobium Chlorophenolicum

ABSTRACT Pentachlorophenol (PCP), a highly toxic anthropogenic pesticide, can be mineralized by Sphingobium chlorophenolicum, a gram-negative bacterium isolated from PCP-contaminated soil. However, degradation of PCP is slow and S. chlorophenolicum cannot tolerate high levels of PCP. We have used genome shuffling to improve the degradation of PCP by S. chlorophenolicum. We have obtained several strains that degrade PCP faster and tolerate higher levels of PCP than the wild-type strain. Several strains obtained after the third round of shuffling can grow on one-quarter-strength tryptic soy broth plates containing 6 to 8 mM PCP, while the original strain cannot grow in the presence of PCP at concentrations higher than 0.6 mM. Some of the mutants are able to completely degrade 3 mM PCP in one-quarter-strength tryptic soy broth, whereas no degradation can be achieved by the wild-type strain. Analysis of several improved strains suggests that the improved phenotypes are due to various combinations of mutations leading to an enhanced growth rate, constitutive expression of the PCP degradation genes, and enhanced resistance to the toxicity of PCP and its metabolites.

Determination of the active site of Sphingobium chlorophenolicum 2,6-dichlorohydroquinone dioxygenase (PcpA)

2009 ◽  
Vol 15 (3) ◽  
pp. 291-301 ◽  
Author(s):  
Timothy E. Machonkin ◽  
Patrick L. Holland ◽  
Kristine N. Smith ◽  
Justin S. Liberman ◽  
Adriana Dinescu ◽  
...  
Keyword(s):  
Active Site ◽  
Sphingobium Chlorophenolicum

scholarly journals S-Glutathionyl-(chloro)hydroquinone reductases: a novel class of glutathione transferases

2010 ◽  
Vol 428 (3) ◽  
pp. 419-427 ◽  
Author(s):  
Luying Xun ◽  
Sara M. Belchik ◽  
Randy Xun ◽  
Yan Huang ◽  
Huina Zhou ◽  
...  
Keyword(s):  
Disulfide Bond ◽  
Distinct Group ◽  
Degradation Pathway ◽  
Glutathione Transferases ◽  
Amino Acid Residues ◽  
Tblastn Search ◽  
Conserved Domain ◽  
Ping Pong ◽  
Sphingobium Chlorophenolicum ◽  
Domain Database

Sphingobium chlorophenolicum completely mineralizes PCP (pentachlorophenol). Two GSTs (glutathione transferases), PcpC and PcpF, are involved in the degradation. PcpC uses GSH to reduce TeCH (tetrachloro-p-hydroquinone) to TriCH (trichloro-p-hydroquinone) and then to DiCH (dichloro-p-hydroquinone) during PCP degradation. However, oxidatively damaged PcpC produces GS-TriCH (S-glutathionyl-TriCH) and GS-DiCH (S-glutathionyl-TriCH) conjugates. PcpF converts the conjugates into TriCH and DiCH, re-entering the degradation pathway. PcpF was further characterized in the present study. It catalysed GSH-dependent reduction of GS-TriCH via a Ping Pong mechanism. First, PcpF reacted with GS-TriCH to release TriCH and formed disulfide bond between its Cys53 residue and the GS moiety. Then, a GSH came in to regenerate PcpF and release GS–SG. A TBLASTN search revealed that PcpF homologues were widely distributed in bacteria, halobacteria (archaea), fungi and plants, and they belonged to ECM4 (extracellular mutant 4) group COG0435 in the conserved domain database. Phylogenetic analysis grouped PcpF and homologues into a distinct group, separated from Omega class GSTs. The two groups shared conserved amino acid residues, for GSH binding, but had different residues for the binding of the second substrate. Several recombinant PcpF homologues and two human Omega class GSTs were produced in Escherichia coli and purified. They had zero or low activities for transferring GSH to standard substrates, but all had reasonable activities for GSH-dependent reduction of disulfide bond (thiol transfer), dehydroascorbate and dimethylarsinate. All the tested PcpF homologues reduced GS-TriCH, but the two Omega class GSTs did not. Thus PcpF homologues were tentatively named S-glutathionyl-(chloro)hydroquinone reductases for catalysing the GSH-dependent reduction of GS-TriCH.

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