Monitoring of proteinase activation in cell apoptosis by capillary electrophoresis with bioengineered fluorescent probe
Monitoring of proteinase activation in cell apoptosis by capillary electrophoresis with bioengineered fluorescent probe
Engineering Village 2
2006 Elsevier Inc.
Accession number: 06229907828
Title: Monitoring of proteinase activation in cell apoptosis by capillary electrophoresis with bioengineered fluorescent probe
Authors: Zhou, Shixia; Lin, Juqiang; Du, Wei; Zhang, Zhihong; Luo, Qingming; Liu, Bi-Feng; Dai, Yiqun
Author affiliation: The Key Laboratory of Biomedical Photonics of MOE, Department of Systems Biology, Huazhong University of Science and Technology, Wuhan 430074, China
Serial title: Analytica Chimica Acta
Abbreviated serial title: Anal. Chim. Acta
Volume: v 569
Issue: n 1-2
Issue date: May 31 2006
Publication year: 2006
Pages: p 176-181
Language: English
ISSN: 0003-2670
CODEN: ACACAM
Document type: Journal article (JA)
Publisher: Elsevier, Amsterdam, 1000 AE, Netherlands
Abstract: In this article, capillary electrophoresis (CE) was demonstrated as a novel means to monitor the activation of proteinase during cell apoptosis. A unique fluorescent probe that fused a specific amino acid sequence DEVD with two fluorescent proteins enhanced cyan fluorescent protein (ECFP) and red fluorescent protein (DsRed), ECFP-DEVD-DsRed, was engineered in HeLa cells as a substrate of proteinase caspase-3. Molecular imaging in vivo was conducted to evaluate the stability and reliability of this fusion protein probe expressed in HeLa cells. With treatment of a certain dose of cisplatin to HeLa cells, apoptosis was initiated, and then caspase-3 was activated that could specifically recognize the DEVD site and would subsequently cleave the constructed fluorescent probe into two pieces of protein fragments. Analyzing the cell lysates with CE in vitro at a series of time points gave a clear description of activation process of caspase-3 in cell apoptosis. Several parameters that influenced CE separation quality were optimized, such as buffer type, pH value, concentration and applied voltage. The employment of two color fluorescent proteins significantly simplified the separation and identification of the residues of the cleavage reaction. ? 2006 Elsevier B.V. All rights reserved.
Number of references: 38
Ei main heading: Enzyme kinetics
Ei controlled terms: Electrophoresis - Chemical activation - Biomedical engineering - Probes - Cells - Proteins - Molecular biology - Imaging techniques
Uncontrolled terms: Capillary electrophoresis - Protein activation - Caspase-3 - Cell apoptosis - Fluorescent proteins
Ei classification codes: 461.8 Biotechnology - 701.1 Electricity: Basic Concepts & Phenomena - 802.2 Chemical Reactions - 461.1 Biomedical Engineering - 941 Acoustical and Optical Measuring Instruments - 461.2 Biological Materials - 804.1 Organic Compounds - 461.9 Biology - 723.2 Data Processing
Treatment: Theoretical (THR); Experimental (EXP)
DOI: 10.1016/j.aca.2006.03.088
Database: Compendex
Compilation and indexing terms, ? 2006 Elsevier Inc. All rights reserved
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