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Monitoring eIF4F Assembly by Measuring eIF4E-eIF4G Interaction in Live Cells
Published on: May 1, 2020
An aptamer-based biosensor for mammalian initiation factor eukaryotic initiation factor 4A.
Akihiro Oguro1, Takashi Ohtsu, Yoshikazu Nakamura
1Department of Basic Medical Sciences, Institute of Medical Science, University of Tokyo, Minato-ku, Tokyo 108-8639, Japan.
Analytical Biochemistry
|March 3, 2009
Summary
Researchers developed an RNA aptamer biosensor to detect eukaryotic initiation factor 4A (eIF4A). This aptamer efficiently purifies and detects eIF4A in cell extracts using affinity pull-down and surface plasmon resonance assays.
Area of Science:
- Biochemistry
- Molecular Biology
- Biotechnology
Background:
- Aptamers are nucleic acid molecules selected for high affinity binding to specific targets.
- Eukaryotic initiation factor 4A (eIF4A) plays a crucial role in protein translation.
- Developing sensitive detection methods for eIF4A is important for biological research.
Purpose of the Study:
- To demonstrate the utility of an RNA aptamer as a biosensor for eIF4A.
- To establish methods for detecting and purifying eIF4A using the selected aptamer.
Main Methods:
- Selection of an RNA aptamer against eIF4A.
- Immobilization of the aptamer to resin for affinity pull-down assays.
- Development of a surface plasmon resonance (SPR) assay using an aptamer-immobilized sensor chip.
Main Results:
- The RNA aptamer effectively purified eIF4A from crude cell extracts.
- Dot-blot analysis detected approximately 300 ng of eIF4A using a 32P-labeled aptamer.
- An optimized SPR assay detected eIF4A at the nanogram level in whole cell lysates in real-time.
Conclusions:
- An RNA aptamer against eIF4A functions as an efficient biosensor.
- The aptamer-based methods enable sensitive detection and purification of eIF4A.
- This work presents a novel real-time sensor for eIF4A in complex biological samples.
