Dataset Information

A twin xanthan lyase-dependent xanthan degradation system in Paenibacillus taichungensis I5

ABSTRACT: Xanthan gum, a natural heteropolysaccharide produced by Xanthomonas species, has many biotechnological applications across industries due to its unique rheological properties. Expanding its utility requires specific enzymes capable of targeted xanthan modification or degradation. In this study, a novel bacterial strain, isolated from a spoiled xanthan sample and identified as Paenibacillus taichungensis I5, was shown to degrade xanthan using a plate screening assay with Congo red. Enzyme activity tests of the culture supernatant demonstrated the secretion of xanthan-degrading enzymes. Genome and proteome analyses suggests a chromosomal xanthan utilization locus encoding a suite of enzymes, including a xanthanase (Pt_XanGH9), two xanthan lyases (Pt_XanPL8a and Pt_XanPL8b), two unsaturated glucuronidases, two α-mannosidases, as well as transport and regulator proteins. Functional characterization through recombinant protein expression and enzyme assays confirmed the functions of Pt_XanGH9, Pt_XanPL8a and Pt_XanPL8b on native xanthan and xanthan-derived oligosaccharides. The polysaccharide degradation products released by these enzymes were identified via LC-MS analysis. The two xanthan lyases differed in cleavage specificity. In contrast to Pt_XanPL8a, Pt_XanPL8b is synthesized with an N-terminal signal peptide, yet both lyases were detected in cell-free supernatant during growth on xanthan. Based on the composition of the xanthan utilization gene cluster and preliminary enzyme characteristics, a working model for xanthan utilization by P. taichungensis I5 is proposed. Reaching a better understanding of bacterial xanthan derivatives and xanthan degrading pathways and the enzymes involved may help to develop modified xanthan derivatives and xanthan degrading enzymes that align with the specific demands of various industrial process.

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ORGANISM(S): Paenibacillus Taichungensis

SUBMITTER: Christina Ludwig

LAB HEAD: Christina Ludwig

PROVIDER: PXD068698 | Pride | 2026-02-02

REPOSITORIES: Pride

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			Action	DRS
	BBM_461_P201_01_FRH_002_R1.raw	Raw
	BBM_461_P201_01_FRH_003_R1.raw	Raw
	BBM_461_P201_01_FRH_004_R1.raw	Raw
	BBM_461_P201_01_FRH_006_R1.raw	Raw
	BBM_461_P201_01_FRH_007_R1.raw	Raw

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A twin xanthan lyase-dependent xanthan degradation system in Paenibacillus taichungensis I5.

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

Applied microbiology and biotechnology 20260122 1

Xanthan gum, a natural heteropolysaccharide produced by Xanthomonas species, has many biotechnological applications across industries due to its unique rheological properties. Expanding its utility requires specific enzymes capable of targeted xanthan modification or degradation. In this study, a novel bacterial strain, isolated from a spoiled xanthan sample and identified as Paenibacillus taichungensis I5, was shown to degrade xanthan using a plate screening assay with Congo red. Activity tests ...[more]

PMID: 41571977

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Project description: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.

Project description:Sugarcane is an economically important crop contributing to the world’s sugar and ethanol production with 80% and 40%, respectively. In recent years, the growing demands for sugar and ethanol production has prompted the necessity to increase sugarcane productivity through conventional breeding programs. However, sugarcane breeders have encountered several difficulties to raise productivity, mainly due to its complex genetics. Sugarcane has a polyploidy genome, with many varieties being aneuploidy. Today, the majority of the planted sugarcane cultivars are complex hybrids derived mainly from crosses between Saccharum officinarum and S. spontaneum. Therefore, proteomics can provide some insight into deciphering gene regulation and changes in carbon metabolism and sucrose accumulation in the culms at different stages of plant development. The aim of this work was to compare the quantitative changes of proteins in sugarcane culms, during plant growth and sucrose accumulation. Total proteins were isolated from both, juvenile and maturing internodes at three stages of plant development. Label free shotgun proteomics was used for protein profiling and quantification. The internodes 5 (I5) and 9 (I9) of 4, 7 and 10 month-old-plants (4M, 7M and 10M, respectively) were harvested and used for proteomic analyses. To mimic field conditions of sucrose accumulation during sugarcane maturation, we stopped watering 10M plants for 10 days. An average of 1130 proteins, unique and differentially expressed across all ages were identified and quantified. Proteins were categorized within 27 functional groups, related to biological process. The patterns of expression for some categories, such as cellular amino acids, metabolic processes, secondary metabolic processes and translation were down-regulated in the immature internode (I5-10M), while up-regulated in the mature I9-10M. We observed an increase in the abundance of several enzymes of the glycolytic pathway and isoforms of alcohol dehydrogenase (ADH) and pyruvate decarboxylase (PDC), in the juvenile stages of development of I9. These changes in enzymes contents indicates that at the early stages of internode development, hypoxia is increasing the glycolytic and ethanolic fermentation pathways, in order to supply ATP for plant growth and NAD+ for mitochondrial respiration, which might be impaired by the low oxygen availability inside the culm.