Characterization of a Novel Peptide from Pathogenic Leptospira and Its Cytotoxic Effect

Paratsaphan, Saksakon; Moonsom, Saengduen; Reamtong, Onrapak; Roytrakul, Sittiruk; Wuthiekanun, Vanaporn; Day, Nicholas P. J.; Sonthayanon, Piengchan

doi:10.3390/pathogens9110906

Cited by 2 publications

(1 citation statement)

References 50 publications

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“…This cell line has been extensively used in the study of several emerging pathogens, due to its high susceptibility to viruses and bacterial toxins, such as diphtheria toxin, heat-labile enterotoxins, and Shiga-like toxins (also known as “verotoxins”) ( Sheets, 2000 ; Naoki et al, 2014 ). In addition, VERO cells have also been used as a model to study potential virulence factors in leptospires ( Lee et al, 2002 ; Lima et al, 2013 ; Chaurasia and Sritharan, 2020 ; Paratsaphan et al, 2020 ). To recover cell-bound phages from VERO cells, we used the BRASIL single-step centrifugation approach.…”

Section: Discussionmentioning

confidence: 99%

Characterization of a virulence-modifying protein of Leptospira interrogans identified by shotgun phage display

et al. 2022

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Pathogenic species of Leptospira are etiologic agents of leptospirosis, an emerging zoonotic disease of worldwide extent and endemic in tropical regions. The growing number of identified leptospiral species sheds light to their genetic diversity and unique virulence mechanisms, many of them still remain unknown. Toxins and adhesins are important virulence factors in several pathogens, constituting promising antigens for the development of vaccines with cross-protection and long-lasting effect against leptospirosis. For this aim, we used the shotgun phage display technique to unravel new proteins with adhesive properties. A shotgun library was constructed using fragmented genomic DNA from Leptospira interrogans serovar Copenhageni strain Fiocruz L1-130 and pG8SAET phagemid vector. Selection of phages bearing new possible cell-binding antigens was performed against VERO cells, using BRASIL biopanning methodology. Analysis of selected clones revealed the hypothetical protein LIC10778, a potentially exposed virulence factor that belongs to the virulence-modifying (VM) protein family (PF07598), composed of 13 members in the leptospiral strain Fiocruz L1-130. Prediction of LIC10778 tertiary structure indicates that the protein contains a cellular-binding domain (N-terminal portion) and an unknown domain of no assigned activity (C-terminal portion). The predicted N-terminal domain shared structural similarities with the cell-binding and internalization domain of toxins like Ricin and Abrin, as well as to the Community-Acquired Respiratory Distress Syndrome (CARDS) toxin in Mycoplasma pneumoniae. Interestingly, recombinant portions of the N-terminal region of LIC10778 protein showed binding to laminin, collagens I and IV, vitronectin, and plasma and cell fibronectins using overlay blotting technique, especially regarding the binding site identified by phage display. These data validate our preliminary phage display biopanning and support the predicted three-dimensional models of LIC10778 protein and other members of PF07598 protein family, confirming the identification of the N-terminal cell-binding domains that are similar to ricin-like toxins. Moreover, fluorescent fused proteins also confirmed that N-terminal region of LIC10778 is capable of binding to VERO and A549 cell lines, further highlighting its virulence role during host-pathogen interaction in leptospirosis probably mediated by its C-terminal domain. Indeed, recent results in the literature confirmed this assumption by demonstrating the cytotoxicity of a closely related PF07598 member.

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Section: Discussionmentioning

confidence: 99%

Characterization of a virulence-modifying protein of Leptospira interrogans identified by shotgun phage display

et al. 2022

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Epitope Analysis of Hypothetical Proteins in Leptospira interrogans Serovar Lai Reveals Potential Diagnostic Markers

Saranya,

Ramya

2024

J. Pure Appl. Microbiol.

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Leptospirosis is a neglected zoonosis caused by a pathogenic spirochete, Leptospira interrogans. The mode of infection in humans is through an abrasion in human skin or the conjunctiva and mucous membrane. Infected patients usually show different symptoms resembling bacterial or viral infections such as the flu. Hence, diagnosing leptospirosis in the early stage is complex, and can be easily confused with other infections. A strategical pathway was developed to analyze the hypothetical proteins in L. interrogans and unveil their potential as diagnostic markers. Subcellular localization tools such as PSORTb, CELLO, SOSUI-GramN, and ProtCompB were used to segregate the outer membrane and surface proteins from the overall pool of hypothetical proteins. The shortlisted proteins were checked for their virulency, and antigenicity through tools such as VirulentPred, and VaxiJen, respectively. Proteins with the highest scores were fed into ElliPro which predicted both linear and discontinuous epitopes in each protein. Proteins with many epitopes were further analyzed with BepiPred 3.0, which provided the epitope probability for each protein’s amino acid. Epitope probability of the potential proteins was compared with the standard diagnostic marker, LipL32. The comparison revealed that a protein (UniProt ID D4YW28) has better immunogenic potential than the gold standard marker, LipL32. In conclusion, this protein can be used as a diagnostic marker for the detection of leptospirosis and it will also serve as a better vaccine candidate.

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Characterization of a Novel Peptide from Pathogenic Leptospira and Its Cytotoxic Effect

Cited by 2 publications

References 50 publications

Characterization of a virulence-modifying protein of Leptospira interrogans identified by shotgun phage display

Characterization of a virulence-modifying protein of Leptospira interrogans identified by shotgun phage display

Epitope Analysis of Hypothetical Proteins in Leptospira interrogans Serovar Lai Reveals Potential Diagnostic Markers

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