Scopus İndeksli Yayınlar Koleksiyonu / Scopus Indexed Publications Collection

Permanent URI for this collectionhttps://hdl.handle.net/11147/7148

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Access Right: Closed AccessAuthor: search.filters.author.Gudlur, Sushanth

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  • Article
    Free-Standing Three-Dimensional Graphene Scaffolds for Protease Functional Assay
    (Elsevier Science Sa, 2024) Ng, Zhi Kai; Yan, Evelias; Goyal, Garima; Gudlur, Sushanth; Kanagavel, Deepankumar; Yildiz, Umit Hakan; Teo, Edwin Hang Tong
    Three-dimensional graphene scaffolds (3d-GS) of high porosity possessing good fluorescence quenching properties are potential candidates for the development of optical biosensors. Herein, we demonstrate the feasibility of utilising intact and free-standing 3d-GS for sensitive detection of proteases, a class of disease diagnosis bio-markers of significant interest. Recombinant OmpT was employed as a model protease for validating the pro-posed methodology. A short (15-residue) peptide sequence encoding a specific recognition site for OmpT was end-labelled with a fluorescent dye (5-FAM) whose fluorescence is quenched when the peptide is anchored to 3d-GS. However, in the presence of OmpT, the peptide is cleaved and released from 3d-GS, resulting in a sig-nificant recovery in fluorescence. The functional assay described herein involves a single step fabrication process of anchoring the peptide to 3d-GS. The integrity of the 3d-GS is hypothesised to overcome the concern of dynamic requenching associated with the typical homogeneous assays based on graphene, yielding a limit of detection (LOD) of similar to 140 nM, which is over an order higher than homogeneous assays performed using the same composition of graphene in powdered form. To the best of our knowledge, this is the first report on utilising free-standing 3d-GS for facile assaying of proteases.
  • Article
    Citation - WoS: 6
    Citation - Scopus: 8
    Outer-Membrane Protease (ompt) Based E.coli Sensing With Anionic Polythiophene and Unlabeled Peptide Substrate
    (John Wiley and Sons Inc., 2020) Sinsinbar, Gaurav; Gudlur, Sushanth; Wood, Sarah E.; Ammanath, Gopal; Yıldız, Ümit Hakan; Alagappan, Palaniappan; Liedberg, Bo
    E. coli and Salmonella are two of the most common bacterial pathogens involved in foodborne and waterborne related deaths. Hence, it is critical to develop rapid and sensitive detection strategies for near-outbreak applications. Reported is a simple and specific assay to detect as low as 1 CFU mL(-1)of E. coli in water within 6 hours by targeting the bacteria's surface protease activity. The assay relies on polythiophene acetic acid (PTAA) as an optical reporter and a short unlabeled peptide (LL37(FRRV)) previously optimized as a substrate for OmpT, an outer-membrane protease on E. coli. LL37(FRRV)interacts with PTAA to enhance its fluorescence while also inducing the formation of a helical PTAA-LL37(FRRV)construct, as confirmed by circular dichroism. However, in the presence of E. coli LL37(FRRV)is cleaved and can no longer affect the conformations and optical properties of PTAA. This ability to distinguish between an intact and cleaved peptide was investigated in detail using LL37(FRRV)sequence variants.