Multiplexed fitness profiling by RB-TnSeq elucidates pathways for lignin-related aromatic catabolism in Sphingobium sp. SYK-6

Alissa Bleem; Ryo Kato; Zoe A Kellermyer; Rui Katahira; Masahiro Miyamoto; Koh Niinuma; Naofumi Kamimura; Eiji Masai; Gregg T Beckham

doi:10.1016/j.celrep.2023.112847

Multiplexed fitness profiling by RB-TnSeq elucidates pathways for lignin-related aromatic catabolism in Sphingobium sp. SYK-6

Cell Rep. 2023 Aug 29;42(8):112847. doi: 10.1016/j.celrep.2023.112847. Epub 2023 Jul 28.

Authors

Alissa Bleem¹, Ryo Kato², Zoe A Kellermyer¹, Rui Katahira¹, Masahiro Miyamoto², Koh Niinuma², Naofumi Kamimura², Eiji Masai³, Gregg T Beckham⁴

Affiliations

¹ Renewable Resources and Enabling Sciences Center, National Renewable Energy Laboratory, Golden, CO 80401, USA.
² Department of Materials Science and Bioengineering, Nagaoka University of Technology, Nagaoka, Niigata 940-2188, Japan.
³ Department of Materials Science and Bioengineering, Nagaoka University of Technology, Nagaoka, Niigata 940-2188, Japan. Electronic address: emasai@vos.nagaokaut.ac.jp.
⁴ Renewable Resources and Enabling Sciences Center, National Renewable Energy Laboratory, Golden, CO 80401, USA. Electronic address: gregg.beckham@nrel.gov.

PMID: 37515767
DOI: 10.1016/j.celrep.2023.112847

Abstract

Bioconversion of lignin-related aromatic compounds relies on robust catabolic pathways in microbes. Sphingobium sp. SYK-6 (SYK-6) is a well-characterized aromatic catabolic organism that has served as a model for microbial lignin conversion, and its utility as a biocatalyst could potentially be further improved by genome-wide metabolic analyses. To this end, we generate a randomly barcoded transposon insertion mutant (RB-TnSeq) library to study gene function in SYK-6. The library is enriched under dozens of enrichment conditions to quantify gene fitness. Several known aromatic catabolic pathways are confirmed, and RB-TnSeq affords additional detail on the genome-wide effects of each enrichment condition. Selected genes are further examined in SYK-6 or Pseudomonas putida KT2440, leading to the identification of new gene functions. The findings from this study further elucidate the metabolism of SYK-6, while also providing targets for future metabolic engineering in this organism or other hosts for the biological valorization of lignin.

Keywords: CP: Microbiology; Pseudomonas putida; Sphingobium sp. SYK-6; Transposon insertion sequencing; acid tolerance; aromatic catabolism; bacterial genetics; lignin.

Publication types

Research Support, U.S. Gov't, Non-P.H.S.
Research Support, Non-U.S. Gov't

MeSH terms

Gene Library
Lignin* / metabolism
Metabolic Engineering*
Secondary Metabolism

Substances

Lignin