Outer-Membrane Protease (OmpT) Based E. coli Sensing with Anionic Polythiophene and Unlabeled Peptide Substrate

Gaurav Sinsinbar; Sushanth Gudlur; Sarah E. Wood; Gopal Ammanath; Hakan U. Yildiz; Palaniappan Alagappan; Milan Mrksich; Bo Liedberg

doi:10.1002/anie.202008444

Outer-Membrane Protease (OmpT) Based E. coli Sensing with Anionic Polythiophene and Unlabeled Peptide Substrate

Gaurav Sinsinbar, Sushanth Gudlur, Sarah E. Wood, Gopal Ammanath, Hakan U. Yildiz, Palaniappan Alagappan, Milan Mrksich, Bo Liedberg^*

^*Corresponding author for this work

Office for Research

Research output: Contribution to journal › Article › peer-review

5 Scopus citations

Abstract

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.

Original language	English (US)
Pages (from-to)	18068-18077
Number of pages	10
Journal	Angewandte Chemie - International Edition
Volume	59
Issue number	41
DOIs	https://doi.org/10.1002/anie.202008444
State	Published - Oct 5 2020

Keywords

enzymes
fluorescence
membrane proteins
pathogens
peptides

ASJC Scopus subject areas

Chemistry(all)
Catalysis

Access to Document

10.1002/anie.202008444

Cite this

@article{cf91d5ee2b6744c1a958fa58c25658c3,

title = "Outer-Membrane Protease (OmpT) Based E. coli Sensing with Anionic Polythiophene and Unlabeled Peptide Substrate",

abstract = "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 (LL37FRRV) previously optimized as a substrate for OmpT, an outer-membrane protease on E. coli. LL37FRRV interacts with PTAA to enhance its fluorescence while also inducing the formation of a helical PTAA-LL37FRRV construct, as confirmed by circular dichroism. However, in the presence of E. coli LL37FRRV 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 LL37FRRV sequence variants.",

keywords = "enzymes, fluorescence, membrane proteins, pathogens, peptides",

author = "Gaurav Sinsinbar and Sushanth Gudlur and Wood, {Sarah E.} and Gopal Ammanath and Yildiz, {Hakan U.} and Palaniappan Alagappan and Milan Mrksich and Bo Liedberg",

note = "Funding Information: This work was funded by the Singapore Ministry of Education Academic Research Fund Tier 2 (MOE2018-T2-1-025) and the NTU-NU Institute for NanoMedicine located at the International Institute for Nanotechnology, Northwestern University, USA and the Nanyang Technological University, Singapore; Agmt10/20/14. We thank Ms. Batika Saxena for help with the graphics. Funding Information: This work was funded by the Singapore Ministry of Education Academic Research Fund Tier 2 (MOE2018‐T2‐1‐025) and the NTU‐NU Institute for NanoMedicine located at the International Institute for Nanotechnology, Northwestern University, USA and the Nanyang Technological University, Singapore; Agmt10/20/14. We thank Ms. Batika Saxena for help with the graphics. Publisher Copyright: {\textcopyright} 2020 Wiley-VCH GmbH",

year = "2020",

month = oct,

day = "5",

doi = "10.1002/anie.202008444",

language = "English (US)",

volume = "59",

pages = "18068--18077",

journal = "Angewandte Chemie - International Edition",

issn = "1433-7851",

publisher = "John Wiley and Sons Ltd",

number = "41",

}

TY - JOUR

T1 - Outer-Membrane Protease (OmpT) Based E. coli Sensing with Anionic Polythiophene and Unlabeled Peptide Substrate

AU - Sinsinbar, Gaurav

AU - Gudlur, Sushanth

AU - Wood, Sarah E.

AU - Ammanath, Gopal

AU - Yildiz, Hakan U.

AU - Alagappan, Palaniappan

AU - Mrksich, Milan

AU - Liedberg, Bo

N1 - Funding Information: This work was funded by the Singapore Ministry of Education Academic Research Fund Tier 2 (MOE2018-T2-1-025) and the NTU-NU Institute for NanoMedicine located at the International Institute for Nanotechnology, Northwestern University, USA and the Nanyang Technological University, Singapore; Agmt10/20/14. We thank Ms. Batika Saxena for help with the graphics. Funding Information: This work was funded by the Singapore Ministry of Education Academic Research Fund Tier 2 (MOE2018‐T2‐1‐025) and the NTU‐NU Institute for NanoMedicine located at the International Institute for Nanotechnology, Northwestern University, USA and the Nanyang Technological University, Singapore; Agmt10/20/14. We thank Ms. Batika Saxena for help with the graphics. Publisher Copyright: © 2020 Wiley-VCH GmbH

PY - 2020/10/5

Y1 - 2020/10/5

N2 - 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 (LL37FRRV) previously optimized as a substrate for OmpT, an outer-membrane protease on E. coli. LL37FRRV interacts with PTAA to enhance its fluorescence while also inducing the formation of a helical PTAA-LL37FRRV construct, as confirmed by circular dichroism. However, in the presence of E. coli LL37FRRV 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 LL37FRRV sequence variants.

AB - 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 (LL37FRRV) previously optimized as a substrate for OmpT, an outer-membrane protease on E. coli. LL37FRRV interacts with PTAA to enhance its fluorescence while also inducing the formation of a helical PTAA-LL37FRRV construct, as confirmed by circular dichroism. However, in the presence of E. coli LL37FRRV 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 LL37FRRV sequence variants.

KW - enzymes

KW - fluorescence

KW - membrane proteins

KW - pathogens

KW - peptides

UR - http://www.scopus.com/inward/record.url?scp=85089495321&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=85089495321&partnerID=8YFLogxK

U2 - 10.1002/anie.202008444

DO - 10.1002/anie.202008444

M3 - Article

C2 - 32618102

AN - SCOPUS:85089495321

SN - 1433-7851

VL - 59

SP - 18068

EP - 18077

JO - Angewandte Chemie - International Edition

JF - Angewandte Chemie - International Edition

IS - 41

ER -

Outer-Membrane Protease (OmpT) Based E. coli Sensing with Anionic Polythiophene and Unlabeled Peptide Substrate

Abstract

Keywords

ASJC Scopus subject areas

Access to Document

Other files and links

Fingerprint

Cite this