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Measurement of Tissue Non-Heme Iron Content using a Bathophenanthroline-Based Colorimetric Assay
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Development of heme protein based oxygen sensing indicators
Jiro Nomata1,2, Toru Hisabori3,4
1Laboratory for Chemistry and Life Science, Institute of Innovative Research, Tokyo Institute of Technology, Nagatsuta 4259, Midori-ku, Yokohama, 226-8503, Japan.
Scientific Reports
|August 9, 2018
Summary
Researchers developed ANA-Y, a sensitive oxygen sensor protein, to monitor oxygen homeostasis in cells. This genetically encoded tool enables real-time measurement of oxygen levels and photosynthetic oxygen production.
Area of Science:
- Biochemistry
- Molecular Biology
- Cell Biology
Background:
- Oxygen is vital for aerobic life, participating in essential biochemical reactions and intracellular signaling.
- Understanding oxygen homeostasis is crucial for studying cellular processes.
- Existing methods for oxygen measurement may have limitations in real-time cellular analysis.
Purpose of the Study:
- To develop a genetically encoded fluorescent protein sensor for real-time monitoring of oxygen levels in living cells.
- To investigate oxygen homeostasis and oxygen-dependent cellular processes.
- To create a sensitive and responsive tool for biological research.
Main Methods:
- Engineered a novel oxygen sensor protein, ANA-Y (Anaerobic/aerobic sensing yellow fluorescence protein).
- Utilized the oxygen sensor domain of bacterial phosphodiesterase direct oxygen sensor protein (DosP) fused to yellow fluorescence protein (YFP) via a coiled-coil linker.
- Characterized ANA-Y's sensitivity, response time, and reversibility.
Main Results:
- ANA-Y demonstrated high sensitivity to oxygen with a half-saturation concentration of 18 μM.
- The sensor exhibited rapid response to dissolved oxygen within 10 seconds.
- ANA-Y enabled direct determination of initial photosynthetic oxygen production in cyanobacteria.
- Reversible fluorescence changes were observed, correlating with oxygen concentration fluctuations.
Conclusions:
- ANA-Y is a robust and sensitive genetically encoded oxygen sensor for monitoring oxygen homeostasis in real-time.
- The sensor's mechanism, involving YFP fluorescence and DosH absorbance changes, can be adapted for developing novel biosensors.
- This tool facilitates the study of oxygen-dependent biological processes, including photosynthesis.
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