Dataset Information

Heterologous expression of clostridithiol genes from Clostridioides difficile in Escherichia coli

ABSTRACT: Detection of clostridithiols from heterologous expression of one or both putative genes for clostridithiol biosynthesis (originating from Clostridioides difficile wild-type 630) in liquid cultures of E. coli BL21(DE3) grown in modified minimal medium

INSTRUMENT(S): 6550 iFunnel Q-TOF LC/MS

ORGANISM(S): Escherichia Coli Bl21(de3) (ncbitaxon:469008)

SUBMITTER: Jason Crawford

PROVIDER: MSV000098169 | MassIVE | Thu Jun 12 10:57:00 BST 2025

REPOSITORIES: MassIVE

ACCESS DATA

Dataset's files

Source:

			Action	DRS
		Other

Items per page:

1 - 1 of 1

Publications

Discovery of a Widespread Polyamine-Low-Molecular-Weight Thiol Hybrid Pathway in Clostridioides difficile.

Hunt Rachelle L RL Oh Joonseok J Jain Abhishek A Kuo Ting-Hao TH Berardi Domenica D Jian Wenya W Song Deguang D Wu Qihao Q Goodman Andrew L AL Palm Noah W NW Zimmermann Michael M Johnson Caroline H CH Crawford Jason M JM

ACS infectious diseases 20250717 8

Clostridioides difficile infection can cause severe inflammation in the gastrointestinal (GI) tract, leading to diarrhea, colitis, and an increased risk of colorectal cancer. Colonization of C. difficile is associated with microbial community-level changes in the expression of polyamine and polyamine precursor biosynthesis genes. Polyamines are abundant cationic metabolites that serve indispensable functions for all kingdoms, particularly in gut homeostasis. Catabolism of the polya ...[more]

PMID: 40671632

Similar Datasets

Project description:Unspecific peroxygenases (UPOs) constitute a new family of fungal heme-thiolate enzymes in which there is high biotechnological interest. Although several thousand genes encoding hypothetical UPO-type proteins have been identified in sequenced fungal genomes and other databases, only a few UPO enzymes have been experimentally characterized to date. Therefore, gene screening and heterologous expression from genetic databases are a priority in the search for ad hoc UPOs for oxyfunctionalization reactions of interest. Very recently, Escherichia coli production of a previously described basidiomycete UPO (as a soluble and active enzyme) has been reported. Here, we explored this convenient heterologous expression system to obtain the protein products from available putative UPO genes. In this way, two UPOs from the ascomycetes Collariella virescens (syn., Chaetomium virescens) and Daldinia caldariorum were successfully obtained, purified, and characterized. Comparison of their kinetic constants for oxidation of model substrates revealed 10- to 20-fold-higher catalytic efficiency of the latter enzyme in oxidizing simple aromatic compounds (such as veratryl alcohol, naphthalene, and benzyl alcohol). Homology molecular models of these enzymes showed three conserved and two differing residues in the distal side of the heme (the latter representing two different positions of a phenylalanine residue). Interestingly, replacement of the C. virescens UPO Phe88 by the homologous residue in the D. caldariorum UPO resulted in an F88L variant with 5- to 21-fold-higher efficiency in oxidizing these aromatic compounds.IMPORTANCE UPOs catalyze regio- and stereoselective oxygenations of both aromatic and aliphatic compounds. Similar reactions were previously described for cytochrome P450 monooxygenases, but UPOs have the noteworthy biotechnological advantage of being stable enzymes requiring only H2O2 to be activated. Both characteristics are related to the extracellular nature of UPOs as secreted proteins. In the present study, the limited repertoire of UPO enzymes available for organic synthesis and other applications is expanded with the description of two new ascomycete UPOs obtained by Escherichia coli expression of the corresponding genes as soluble and active enzymes. Moreover, directed mutagenesis in E. coli, together with enzyme molecular modeling, provided relevant structure-function information on aromatic substrate oxidation by these two new biocatalysts.

Dataset Information

Heterologous expression of clostridithiol genes from Clostridioides difficile in Escherichia coli

Dataset's files

Publications

Discovery of a Widespread Polyamine-Low-Molecular-Weight Thiol Hybrid Pathway in <i>Clostridioides difficile</i>.

Similar Datasets

OmicsDI is part of the ELIXIR infrastructure

Tweets

Dataset Information

Heterologous expression of clostridithiol genes from Clostridioides difficile in Escherichia coli

Dataset's files

Publications

Discovery of a Widespread Polyamine-Low-Molecular-Weight Thiol Hybrid Pathway in &lt;i&gt;Clostridioides difficile&lt;/i&gt;.

Similar Datasets

OmicsDI is part of the ELIXIR infrastructure

Tweets

Discovery of a Widespread Polyamine-Low-Molecular-Weight Thiol Hybrid Pathway in <i>Clostridioides difficile</i>.