Optical Fingerprints of Proteases and Their Inhibited Complexes Provided by Differential Cross-Reactivity of Fluorophore-Labeled Single-Stranded DNA
- Shunsuke Tomita*Shunsuke Tomita*E-mail: [email protected]Biomedical Research Institute, National Institute of Advanced Industrial Science and Technology, 1-1-1 Higashi, Tsukuba, Ibaraki 305-8566, JapanDBT-AIST International Laboratory for Advanced Biomedicine (DAILAB), DBT-AIST International Center for Translational & Environmental Research (DAICENTER), National Institute of Advanced Industrial Science and Technology, 1-1-1 Higashi, Tsukuba, Ibaraki 305-8566, JapanMore by Shunsuke Tomita,
- Hiroka SugaiHiroka SugaiBiomedical Research Institute, National Institute of Advanced Industrial Science and Technology, 1-1-1 Higashi, Tsukuba, Ibaraki 305-8566, JapanMore by Hiroka Sugai,
- Masahiro MimuraMasahiro MimuraBiomedical Research Institute, National Institute of Advanced Industrial Science and Technology, 1-1-1 Higashi, Tsukuba, Ibaraki 305-8566, JapanFaculty of Pure and Applied Sciences, University of Tsukuba, 1-1-1 Tennodai, Tsukuba, Ibaraki 305-8573, JapanMore by Masahiro Mimura,
- Sayaka IshiharaSayaka IshiharaBiomedical Research Institute, National Institute of Advanced Industrial Science and Technology, 1-1-1 Higashi, Tsukuba, Ibaraki 305-8566, JapanMore by Sayaka Ishihara,
- Kentaro ShirakiKentaro ShirakiFaculty of Pure and Applied Sciences, University of Tsukuba, 1-1-1 Tennodai, Tsukuba, Ibaraki 305-8573, JapanMore by Kentaro Shiraki, and
- Ryoji Kurita*Ryoji Kurita*E-mail: [email protected]Biomedical Research Institute, National Institute of Advanced Industrial Science and Technology, 1-1-1 Higashi, Tsukuba, Ibaraki 305-8566, JapanDBT-AIST International Laboratory for Advanced Biomedicine (DAILAB), DBT-AIST International Center for Translational & Environmental Research (DAICENTER), National Institute of Advanced Industrial Science and Technology, 1-1-1 Higashi, Tsukuba, Ibaraki 305-8566, JapanFaculty of Pure and Applied Sciences, University of Tsukuba, 1-1-1 Tennodai, Tsukuba, Ibaraki 305-8573, JapanMore by Ryoji Kurita
Abstract
The detection of proteases and their complexes with inhibitor proteins is of great importance for diagnosis and medical-treatment applications. In this study, we report a fingerprint-based sensor using an array of single-stranded DNAs (ssDNAs) labeled with environment-responsive 3′-carboxytetramethylrhodamine (TAMRA) for the identification of proteases. Four TAMRA-modified ssDNAs with different sequences solubilized in two different buffer solutions were incorporated in an array that was capable of generating fluorescent fingerprints unique to the proteases through diverse cross-reactive interactions, allowing the discrimination of (i) 8 proteases and (ii) 12 different mixtures of trypsin and its inhibitor protein (α1-antitrypsin) by multivariate analysis. Constructing an array with TAMRA-modified DNA aptamers that bind to different sites of human thrombin provides fluorescence fingerprints that reflect a reduction of the exposed surface area of thrombin upon complexation with antithrombin III, even in the presence of human serum. We finally demonstrate the potential of hybridization with complementary DNAs as an effective means to easily double the fingerprint information for proteases. Our approach based on the cross-reactive capability of ssDNAs enables high-throughput fingerprint-based sensing that can be flexibly designed and easily constructed, not only for the identification of a variety of proteins including proteases but also for the evaluation of their complexation ability.
Cited By
This article is cited by 3 publications.
- Shunsuke Tomita, Hiroka Sugai, Sayaka Ishihara, Takuya Hosokai, Ryoji Kurita. A Biomimetic Sensor Array Based on a Single Fluorescent Block-copolymer for the Pattern Recognition of Proteins. Chemistry Letters 2020, 49 (12) , 1447-1451. https://doi.org/10.1246/cl.200438
- Shunsuke TOMITA, Hiroka SUGAI, Tsukuru MINAMIKI, Ryoji KURITA. Molecular array device and multivariate analysis for biological fluids. Denki Kagaku 2020, 88 (3) , 262-271. https://doi.org/10.5796/denkikagaku.20-TE0006
- Mari Okada, Hiroka Sugai, Shunsuke Tomita, Ryoji Kurita. A Multichannel Pattern-Recognition-Based Protein Sensor with a Fluorophore-Conjugated Single-Stranded DNA Set. Sensors 2020, 20 (18) , 5110. https://doi.org/10.3390/s20185110