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Title: Rapid, parallel identification of pathways for catabolism of lignin-derived aromatic compounds in Novosphingobium aromaticivorans

Journal Article · · Applied and Environmental Microbiology
DOI:https://doi.org/10.1128/AEM.01185-18· OSTI ID:1471938
 [1];  [2]; ORCiD logo [3]; ORCiD logo [3]
  1. Oak Ridge National Lab. (ORNL), Oak Ridge, TN (United States); Univ. of Arizona, Tucson, AZ (United States)
  2. Oak Ridge National Lab. (ORNL), Oak Ridge, TN (United States); Univ. of Tennessee, Knoxville, TN (United States)
  3. Oak Ridge National Lab. (ORNL), Oak Ridge, TN (United States)

Transposon mutagenesis is a powerful technique in microbial genetics for the identification of genes in uncharacterized pathways. Recently, the throughput of transposon mutagenesis techniques has been dramatically increased through the combination of DNA barcoding and high-throughput sequencing. Here we show that, when applied to catabolic pathways, barcoded transposon libraries can be used to distinguish redundant pathways, decompose complex pathways into substituent modules, discriminate between enzyme homologs, and rapidly identify previously-hypothetical enzymes in an unbiased genome-scale search. We use this technique to identify two genes, which we name desC and desD, are involved in the degradation of the lignin-derived aromatic compound sinapic acid in the non-model bacterium Novosphingobium aromaticivorans. We show that DesC is a methyl-esterase acting on an intermediate formed during sinapic acid catabolism, providing the last enzyme in a proposed catabolic pathway. Here, this approach will be particularly useful in the identification of complete pathways suitable for heterologous expression in metabolic engineering.

Research Organization:
Oak Ridge National Lab. (ORNL), Oak Ridge, TN (United States)
Sponsoring Organization:
USDOE
Grant/Contract Number:
AC05-00OR22725
OSTI ID:
1471938
Journal Information:
Applied and Environmental Microbiology, Vol. 84, Issue 22; ISSN 0099-2240
Publisher:
American Society for MicrobiologyCopyright Statement
Country of Publication:
United States
Language:
English
Citation Metrics:
Cited by: 24 works
Citation information provided by
Web of Science

References (38)

Aromatic-degrading Sphingomonas isolates from the deep subsurface. journal January 1995
A corrinoid-dependent catabolic pathway for growth of a Methylobacterium strain with chloromethane journal April 1999
Comparative genomic analysis reveals distinct genotypic features of the emerging pathogen Haemophilus influenzae type f journal January 2014
Pathways for degradation of lignin in bacteria and fungi journal January 2011
Complete Sequence of a 184-Kilobase Catabolic Plasmid from Sphingomonas aromaticivorans F199 journal March 1999
Unravelling the gallic acid degradation pathway in bacteria: the gal cluster from Pseudomonas putida: Aerobic gallic acid degradation journal November 2010
Markerless Gene Deletion System for Sphingomonads journal March 2012
Structural and Kinetic Characterization of the 4-Carboxy-2-hydroxymuconate Hydratase from the Gallate and Protocatechuate 4,5-Cleavage Pathways of Pseudomonas putida KT2440 journal February 2016
Development of an Optimized Medium, Strain and High-Throughput Culturing Methods for Methylobacterium extorquens journal April 2013
Identifying Genetic Determinants Needed to Establish a Human Gut Symbiont in Its Habitat journal September 2009
A bacterial aromatic aldehyde dehydrogenase critical for the efficient catabolism of syringaldehyde journal March 2017
Ecology, physiology, and phylogeny of deep subsurface Sphingomonas sp journal October 1999
The TraDIS toolkit: sequencing and analysis for dense transposon mutant libraries journal January 2016
Bacterial catabolism of lignin-derived aromatics: New findings in a recent decade: Update on bacterial lignin catabolism: Bacterial catabolism of lignin-derived aromatics journal November 2017
Simultaneous identification of bacterial virulence genes by negative selection journal July 1995
Replica Plating and Indirect Selection of Bacterial Mutants journal January 1952
Tn-seq: high-throughput parallel sequencing for fitness and genetic interaction studies in microorganisms journal September 2009
The art and design of genetic screens: Escherichia coli journal June 2003
Mutagenesis by insertion of a drug-resistance element carrying an inverted repetition journal October 1975
Simultaneous assay of every Salmonella Typhi gene using one million transposon mutants journal October 2009
Chloromethane-Induced Genes Define a Third C1 Utilization Pathway in Methylobacterium chloromethanicum CM4 journal July 2002
Genetic and Biochemical Investigations on Bacterial Catabolic Pathways for Lignin-Derived Aromatic Compounds journal January 2007
Transfer of a Catabolic Pathway for Chloromethane in Methylobacterium Strains Highlights Different Limitations for Growth with Chloromethane or with Dichloromethane journal July 2016
Tracking insertion mutants within libraries by deep sequencing and a genome-wide screen for Haemophilus genes required in the lung journal September 2009
Unusual transformations in the biosynthesis of the antibiotic phosphinothricin tripeptide journal July 2007
Engineering Cellular Metabolism journal March 2016
Molecular characteristics of xenobiotic-degrading sphingomonads journal November 2008
Cross-biome metagenomic analyses of soil microbial communities and their functional attributes journal December 2012
Degradation of 3- O -methylgallate in Sphingomonas paucimobilis SYK-6 by pathways involving protocatechuate 4,5-dioxygenase journal September 2007
Filling gaps in bacterial amino acid biosynthesis pathways with high-throughput genetics journal January 2018
The Homogentisate Pathway: a Central Catabolic Pathway Involved in the Degradation of L-Phenylalanine, L-Tyrosine, and 3-Hydroxyphenylacetate in Pseudomonas putida journal July 2004
Rapid Quantification of Mutant Fitness in Diverse Bacteria by Sequencing Randomly Bar-Coded Transposons journal May 2015
Identification of Three Alcohol Dehydrogenase Genes Involved in the Stereospecific Catabolism of Arylglycerol- -Aryl Ether by Sphingobium sp. Strain SYK-6 journal June 2009
A Tetrahydrofolate-Dependent O-Demethylase, LigM, Is Crucial for Catabolism of Vanillate and Syringate in Sphingomonas paucimobilis SYK-6 journal March 2005
A Novel Tetrahydrofolate-Dependent O-Demethylase Gene Is Essential for Growth of Sphingomonas paucimobilis SYK-6 with Syringate journal April 2004
Characterization of the Gallate Dioxygenase Gene: Three Distinct Ring Cleavage Dioxygenases Are Involved in Syringate Degradation by Sphingomonas paucimobilis SYK-6 journal July 2005
Characterization of the 3-O-Methylgallate Dioxygenase Gene and Evidence of Multiple 3-O-Methylgallate Catabolic Pathways in Sphingomonas paucimobilis SYK-6 journal July 2004
‘Unknown’ proteins and ‘orphan’ enzymes: the missing half of the engineering parts list – and how to find it journal December 2009

Cited By (3)

Isolation of a novel platform bacterium for lignin valorization and its application in glucose-free cis,cis-muconate production journal May 2019
Horizontal transfer of a pathway for coumarate catabolism unexpectedly inhibits purine nucleotide biosynthesis journal October 2019
Modular Engineering of Biomass Degradation Pathways journal April 2019

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