Dataset Information

Identification of a novel xanthan-binding module of a multi-modular Cohnella sp. xanthanase

ABSTRACT: A new strain of xanthan-degrading bacteria identified as Cohnella sp. has been isolated from a xanthan thickener for food production. The strain was able to utilize xanthan as the only carbon source and reduce the viscosity of the xanthan-containing medium during cultivation. Xanthan hydrolytic activity was revealed by congo red staining after growth of the strain on agar plates containing xanthan. By analyzing the secretome of Cohnella sp. after growth on different media (lysogeny broth, glucose mineral medium, xanthan mineral medium), a xanthanase designated as CspXan9 was found and its gene was successfully expressed in Escherichia coli Rosetta2. CspXan9 could efficiently degrade the -1,4-glucan backbone of xanthan after previous removal of pyruvylated mannose residues from the ends of the native xanthan side chains by xanthan lyase treatment (XLT-xanthan). Compared with a known xanthanase from Paenibacillus nanensis, the modular xanthanase CspXan9 had a different module composition at the N- and C-terminal ends. High-performance anion-exchange chromatography (HPAEC-PAD) analysis revealed that the main putative end products released from XLT-xanthan by CspXan9 hydrolysis were tetrasaccharides. Deletion derivatives lacking some of the non-catalytic domains (CspXan9-C, CspXan9-N, CspXan9-C-N) were produced in E. coli to explore the functions of the N- and C-terminal regions of the enzyme. Enzyme assays with the purified deletion derivatives, which all contained the catalytic glycoside hydrolase family 9 (GH9) module, resulted in a range of specific activities on XLT-xanthan between 10.31 ± 0.29 U/mg (CspXan9-C) and 1.38 ± 0.05 U/mg (CspXan9-C-N). Mobility shift assays performed by native affinity polyacrylamide gel electrophoresis (NAPAGE) in the presence of different polysaccharides indicated that the C-terminal module of CspXan9 represents a novel carbohydrate-binding module of CBM66 with binding affinity for XLT-xanthan. The only previously known binding function of a member of the CBM66 family is exo-type binding to the non-reducing fructose ends of the -fructan polysaccharides inulin and levan.

INSTRUMENT(S): Q Exactive HF

ORGANISM(S): Cohnella Sp.

SUBMITTER: Christina Ludwig

LAB HEAD: Christina Ludwig

PROVIDER: PXD050379 | Pride | 2024-06-14

REPOSITORIES: Pride

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Dataset's files

Source:

			Action	DRS
	BBM_461_P201_01_FRH_001.raw	Raw
	BBM_461_P201_01_FRH_005_R1.raw	Raw
	BBM_461_P201_01_FRH_009_R1.raw	Raw
	BBM_461_P201_01_FRH_010_R1.raw	Raw
	BBM_461_P201_01_FRH_011_R1.raw	Raw

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Publications

Identification of a novel xanthan-binding module of a multi-modular Cohnella sp. xanthanase.

Han Rui R Baudrexl Melanie M Ludwig Christina C Berezina Oksana V OV Rykov Sergey V SV Liebl Wolfgang W

Frontiers in microbiology 20240326

A new strain of xanthan-degrading bacteria identified as Cohnella sp. has been isolated from a xanthan thickener for food production. The strain was able to utilize xanthan as the only carbon source and to reduce the viscosity of xanthan-containing medium during cultivation. Comparative analysis of the secretomes of Cohnella sp. after growth on different media led to the identification of a xanthanase designated as CspXan9, which was isolated after recombinant production in ...[more]

PMID: 38596379

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Project description:Arabidopsis thaliana transcriptome analysis in response to plant growth promoting rhizobacteria (PGPR) Experiment 1 : Changes in gene expression profile triggered during root architecture response to Phyllobacterium. Biological question : Which genes are up- or down-regulated in Arabidopsis thaliana cultivated in vitro with increased lateral root development in response to Phyllobacterium STM196 inoculation. Experiment description: Seeds of wild-type Arabidopsis thaliana (ecotype Columbia) were surface-sterilized and sown on agar mineral medium (see below). 4 days after storage in the dark at 4C, seedling were cultivated 6 days in a growth chamber (16 h daily, 20-22C) and then transferred on a fresh agar mineral medium inoculated or not with Phyllobacterium STM196 (2.108 cfu/ml). 6 days later, root and leaves were collected, froze on liquid nitrogen and stored at -80C. Experiment 2 : Changes in gene expression profile triggered during induced systemic resistance (ISR) Biological question : Which genes are up- or down-regulated during the ISR triggered by a rhizobacteria, in comparison with those affected by a pathogenic interaction. Experiment description: Seeds were sown on 0.8% (W/V) agar mineral medium (see below). 4 days after storage in the dark at 4C, seedling were cultivated 6 days in a growth chamber (16 h daily, 20-22C) and then transferred on soil inoculated or not with 107 cfu.g-1 of Bradyrhizobium strain ORS278. Three weeks later, 3 leaves per plant were infiltrated with a suspension of Pseudomonas syringae pv. tomato (2.105 cfu.ml-1) or with MgSO4 10 mM alone for control plants. Infiltrated leaves were collected 24h later. Experiment 3 : Comparison of the effects of 3 rhizobacteria on Arabidopsis thaliana transcriptome Biological question : which genes are specifically induced or repressed in Arabidopsis thaliana by inoculation of the soil with a PGPR vs a bacteria that has the ability to trigger nodule formation in a Legume. Experiment description: Seeds of wild-type Arabidopsis thaliana (ecotype Columbia) were surface-sterilized and sown on agar mineral medium. Four days after storage in the dark at 4C, seedlings were cultivated 6 days in a growth chamber (16 h daily, 20-22C) and then transferred on soil inoculated or not with 108 cfu.g-1 of Mesorhizobium loti, or 108 cfu.g-1 of Phyllobacterium STM196, or 107 cfu.g-1 of Bradyrhizobium ORS278.

Dataset Information

Identification of a novel xanthan-binding module of a multi-modular Cohnella sp. xanthanase

Dataset's files

Publications

Identification of a novel xanthan-binding module of a multi-modular <i>Cohnella</i> sp. xanthanase.

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