Phase-separating peptides recruiting aggregation-induced emission fluorogen for rapid E. coli detection

Rationally designed biomolecular condensates have found applications primarily as drug-delivery systems, thanks to their ability to self-assemble under physico-chemical triggers (such as temperature, pH, or ionic strength) and to concomitantly trap client molecules with exceptionally high efficiency...

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Main Authors: Maricar, Syed, Gudlur, Sushanth, Miserez, Ali
Other Authors: School of Materials Science and Engineering
Format: Article
Language:English
Published: 2023
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Online Access:https://hdl.handle.net/10356/170395
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Institution: Nanyang Technological University
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spelling sg-ntu-dr.10356-1703952023-09-15T15:43:31Z Phase-separating peptides recruiting aggregation-induced emission fluorogen for rapid E. coli detection Maricar, Syed Gudlur, Sushanth Miserez, Ali School of Materials Science and Engineering Center for Sustainable Materials (SusMat) Science::Chemistry::Analytical chemistry Science::Chemistry::Analytical chemistry::Qualitative analysis Aggregation-Induced Emissions Bio-Molecular Rationally designed biomolecular condensates have found applications primarily as drug-delivery systems, thanks to their ability to self-assemble under physico-chemical triggers (such as temperature, pH, or ionic strength) and to concomitantly trap client molecules with exceptionally high efficiency (>99%). However, their potential in (bio)sensing applications remains unexplored. Here, we describe a simple and rapid assay to detect E. coli by combining phase-separating peptide condensates containing a protease recognition site, within which an aggregation-induced emission (AIE)-fluorogen is recruited. The recruited AIE-fluorogen’s fluorescence is easily detected with the naked eye when the samples are viewed under UV-A light. In the presence of E. coli, the bacteria’s outer membrane protease (OmpT) cleaves the phase-separating peptides at the encoded protease recognition site, resulting in two shorter peptide fragments incapable of liquid-liquid phase separation. As a result, no condensates are formed and the fluorogen remains non-fluorescent. The assay feasibility was first tested with recombinant OmpT reconstituted in detergent micelles and subsequently confirmed with E. coli K-12. In its current format, the assay can detect E. coli K-12 (10^8 CFU) within 2 h in spiked water samples and 1-10 CFU/mL with the addition of a 6-7 h pre-culture step. In comparison, most commercially available E. coli detection kits can take anywhere from 8 to 24 h to report their results. Optimizing the peptides for OmpT’s catalytic activity can significantly improve the detection limit and assay time. Besides detecting E. coli, the assay can be adapted to detect other Gram-negative bacteria as well as proteases having diagnostic relevance. Ministry of Education (MOE) Submitted/Accepted version This research was funded by the Ministry of Education (MOE), Singapore, through an Academic Research Fund (AcRF) Tier 3 grant (GrantNo. MOE2019-T3-1-012). 2023-09-13T02:18:10Z 2023-09-13T02:18:10Z 2023 Journal Article Maricar, S., Gudlur, S. & Miserez, A. (2023). Phase-separating peptides recruiting aggregation-induced emission fluorogen for rapid E. coli detection. Analytical Chemistry, 95(26), 9924-9931. https://dx.doi.org/10.1021/acs.analchem.3c01046 0003-2700 https://hdl.handle.net/10356/170395 10.1021/acs.analchem.3c01046 2-s2.0-85165122351 26 95 9924 9931 en MOE 2019-T3-1-012 Analytical Chemistry This document is the Accepted Manuscript version of a Published Work that appeared in final form in Analytical Chemistry, copyright © 2023 American Chemical Society, after peer review and technical editing by the publisher. To access the final edited and published work see https://doi.org/10.1021/acs.analchem.3c01046. application/pdf application/pdf
institution Nanyang Technological University
building NTU Library
continent Asia
country Singapore
Singapore
content_provider NTU Library
collection DR-NTU
language English
topic Science::Chemistry::Analytical chemistry
Science::Chemistry::Analytical chemistry::Qualitative analysis
Aggregation-Induced Emissions
Bio-Molecular
spellingShingle Science::Chemistry::Analytical chemistry
Science::Chemistry::Analytical chemistry::Qualitative analysis
Aggregation-Induced Emissions
Bio-Molecular
Maricar, Syed
Gudlur, Sushanth
Miserez, Ali
Phase-separating peptides recruiting aggregation-induced emission fluorogen for rapid E. coli detection
description Rationally designed biomolecular condensates have found applications primarily as drug-delivery systems, thanks to their ability to self-assemble under physico-chemical triggers (such as temperature, pH, or ionic strength) and to concomitantly trap client molecules with exceptionally high efficiency (>99%). However, their potential in (bio)sensing applications remains unexplored. Here, we describe a simple and rapid assay to detect E. coli by combining phase-separating peptide condensates containing a protease recognition site, within which an aggregation-induced emission (AIE)-fluorogen is recruited. The recruited AIE-fluorogen’s fluorescence is easily detected with the naked eye when the samples are viewed under UV-A light. In the presence of E. coli, the bacteria’s outer membrane protease (OmpT) cleaves the phase-separating peptides at the encoded protease recognition site, resulting in two shorter peptide fragments incapable of liquid-liquid phase separation. As a result, no condensates are formed and the fluorogen remains non-fluorescent. The assay feasibility was first tested with recombinant OmpT reconstituted in detergent micelles and subsequently confirmed with E. coli K-12. In its current format, the assay can detect E. coli K-12 (10^8 CFU) within 2 h in spiked water samples and 1-10 CFU/mL with the addition of a 6-7 h pre-culture step. In comparison, most commercially available E. coli detection kits can take anywhere from 8 to 24 h to report their results. Optimizing the peptides for OmpT’s catalytic activity can significantly improve the detection limit and assay time. Besides detecting E. coli, the assay can be adapted to detect other Gram-negative bacteria as well as proteases having diagnostic relevance.
author2 School of Materials Science and Engineering
author_facet School of Materials Science and Engineering
Maricar, Syed
Gudlur, Sushanth
Miserez, Ali
format Article
author Maricar, Syed
Gudlur, Sushanth
Miserez, Ali
author_sort Maricar, Syed
title Phase-separating peptides recruiting aggregation-induced emission fluorogen for rapid E. coli detection
title_short Phase-separating peptides recruiting aggregation-induced emission fluorogen for rapid E. coli detection
title_full Phase-separating peptides recruiting aggregation-induced emission fluorogen for rapid E. coli detection
title_fullStr Phase-separating peptides recruiting aggregation-induced emission fluorogen for rapid E. coli detection
title_full_unstemmed Phase-separating peptides recruiting aggregation-induced emission fluorogen for rapid E. coli detection
title_sort phase-separating peptides recruiting aggregation-induced emission fluorogen for rapid e. coli detection
publishDate 2023
url https://hdl.handle.net/10356/170395
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