Autotransporter-based surface expression and complementation of split TreA fragments utilized for the detection of antibodies against bovine leukemia virus

Mukherjee, S. K.; Buck, Jeroen De

doi:10.1016/j.jim.2021.113084

Cited by 7 publications

(4 citation statements)

References 19 publications

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“…SpoIIID was selected as the DNA-binding protein of choice because its binding to DNA as a monomer is critical with regards to avoiding self-complementation of the fusion proteins (30). According to multiple studies, split TreA is capable of protein complementation by recombination of the TreA fragments fused to analyte-sensing elements (25)(26)(27)(28). Based on these characteristics, the combination of split TreA platform and SpoIIID can be used to detect the amplicons incorporated with tandem SpoIIID specific recognition sites, indicating presence or absence through the glucose signals generated by the conditional recovery of the trehalase capacity.…”

Section: Discussionmentioning

confidence: 99%

“…Glucose, the main product of this restoration, can be quantified using a colorimetric assay or a handheld glucometer ( 25 ). Various diagnostic assays utilize split TreA fusion proteins to detect biomarkers, including immunoglobulin G, blood calcium, and anti-bovine leukemia virus antibodies ( 26 – 28 ). By incorporating a specific protein-binding sequence into PCR primers, the Gram-types of mastitis pathogens were discriminated in previous research ( 29 ).…”

Section: Introductionmentioning

confidence: 99%

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Discriminating bovine mastitis pathogens by combining loop-mediated isothermal amplification and amplicon-binding split trehalase assay

Miao,

De Buck

2024

Front. Vet. Sci.

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Bovine mastitis is predominantly caused by intramammary infections with various Gram-positive and Gram-negative bacteria, requiring accurate pathogen identification for effective treatment and antimicrobial resistance prevention. Here, a novel diagnostic method was developed to detect mastitis pathogens in milk samples by combining loop-mediated isothermal amplification with a split enzyme biosensor whereby trehalase fragments were fused with a DNA-binding protein, SpoIIID. Three primer sets, LAMPstaph, LAMPstrep, and LAMPneg, harboring SpoIIID recognition sequences targeted Staphylococcus, Streptococcus, and Gram-negative pathogens, respectively. Limits of detection were determined for DNA extracted from bacterial culture and bacteria-spiked milk. The combined method detected as low as 2, 24, and 10 copies of genomic DNA of staphylococci, streptococci and Escherichia coli and 11 CFU/ml for milk spiked with Escherichia coli. Higher detection limits were observed for Gram-positive bacteria in spiked milk. When testing genomic DNA of 10 mastitis isolates at concentrations of 106 and 103 copies per reaction, no cross-reactivity was detected for LAMPstaph nor LAMPstrep, whereas the LAMPneg assay cross-reacted only with Corynebacterium sp. at the highest concentration. This combined method demonstrated the potential to distinguish mastitis pathogenic Gram types for a rapid decision of antimicrobial treatment without culturing.

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Section: Discussionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Discriminating bovine mastitis pathogens by combining loop-mediated isothermal amplification and amplicon-binding split trehalase assay

Miao,

De Buck

2024

Front. Vet. Sci.

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“…The study explored the application of split enzyme sensor diagnostic technology to identify immunoglobulins and antigen-specific antibodies using Escherichia coli str. K-12 periplasmic TreA ( Mukherjee and De Buck 2021 ) ( Fig. 6A ).…”

Section: Application Of Trehalose-degrading Enzymes and Their Future ...mentioning

confidence: 99%

Elucidation of bacterial trehalose-degrading trehalase and trehalose phosphorylase: physiological significance and its potential applications

Shrestha,

Karmacharya,

Han

et al. 2023

Glycobiology

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Bacteria possess diverse metabolic and genetic processes, resulting in the inability of certain bacteria to degrade trehalose. However, some bacteria do have the capability to degrade trehalose, utilizing it as a carbon source, and for defense against environmental stress. Trehalose, a disaccharide, serves as a carbon source for many bacteria, including some that are vital for pathogens. The degradation of trehalose is carried out by enzymes like trehalase (EC 3.2.1.28) and trehalose phosphorylase (EC 2.4.1.64/2.4.1.231), which are classified under the glycoside hydrolase families GH37, GH15, and GH65. Numerous studies and reports have explored the physiological functions, recombinant expression, enzymatic characteristics, and potential applications of these enzymes. However, further research is still being conducted to understand their roles in bacteria. This review aims to provide a comprehensive summary of the current understanding of trehalose degradation pathways in various bacteria, focusing on three key areas: (i) identifying different trehalose-degrading enzymes in Gram-positive and Gram-negative bacteria, (ii) elucidating the mechanisms employed by trehalose-degrading enzymes belonging to the glycoside hydrolases GH37, GH15, and GH65, and (iii) discussing the potential applications of these enzymes in different sectors. Notably, this review emphasizes the bacterial trehalose-degrading enzymes, specifically trehalases (GH37, GH15, and GH65) and trehalose phosphorylases (GH65), in both Gram-positive and Gram-negative bacteria, an aspect that has not been highlighted before.

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“…Recently, a new type of biosensor platform has been developed. Based on an existing blood glucose meter (one of the most successful biosensors for monitoring blood glucose concentration in the market), using E. coli periplasmic trehalase treA as a new lyase, which can convert a variety of analytes into glucose, increasing the versatility of detection even antibodies [108], whole pathogens, protein-protein binding, and protein aggregation [109,110]. Palazzo et al developed a novel optical biosensor for the determination of trehalose levels by immobilizing trehalase, glucose oxidase and horseradish peroxidase layer by layer on a transparent carrier to catalyze the cascade reaction in a series [111] with limit of detection of 10 µM, and linear response of up to 4 mM.…”

Section: Biosensor Developmentmentioning

confidence: 99%

Characterization, heterologous expression and engineering of trehalase for biotechnological applications

Gao

Gong

et al. 2022

Syst Microbiol and Biomanuf

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Trehalose is a non-reducing disaccharide connected by α-1,1-glycosidic bonds; it is widely distributed in bacteria, fungi, yeast, insects, and plant tissues and plays various roles. It can be hydrolyzed by trehalase into two glucose molecules. Trehalases from different sources have been expressed in Escherichia coli, Pichia pastoris, Saccharomyces cerevisiae, baculovirus-silkworm, and other expression systems; however, it is most common in E. coli. The structural characteristics of different glycoside hydrolase (GH) family trehalases and the sources of trehalase have been analyzed. The catalytic mechanism of GH37 trehalase has also been elucidated in detail. Moreover, the molecular modification of trehalase has mainly focused on directed evolution for improving enzyme activity. We comprehensively reviewed the current application status and adaptable transformations was comprehensively overviewed in the context of industrial performance. We suggest that the level of recombinant production is far from meeting industrial requirements, and the catalytic performance of trehalase needs to be improved urgently. Finally, we discuss developmental prospects and future trends.

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Autotransporter-based surface expression and complementation of split TreA fragments utilized for the detection of antibodies against bovine leukemia virus

Cited by 7 publications

References 19 publications

Discriminating bovine mastitis pathogens by combining loop-mediated isothermal amplification and amplicon-binding split trehalase assay

Discriminating bovine mastitis pathogens by combining loop-mediated isothermal amplification and amplicon-binding split trehalase assay

Elucidation of bacterial trehalose-degrading trehalase and trehalose phosphorylase: physiological significance and its potential applications

Characterization, heterologous expression and engineering of trehalase for biotechnological applications

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