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Published on: January 5, 2018
[Isolation of the alternative oxidase from Arum maculatum]
1Xiamen Medical College, Xiamen 361008, China. wangyaying@vip.sina.com
Abstract:
There is plenty of alternate oxidase (AOX) in the inflorescences of thermogenic A rum maculatum. The isolated mitochondria exhibited a high activity, consuming oxygen on average 32 micromoles/min. The concentration of the isolated protein from mitochondria was 14.0 mg/ml. The isolated mitochondria were crashed by osmotic method to isolate matrix, membrane, and membrane soluble and insoluble proteins. The whole mitochondria, membrane and membrane soluble protein showed AOX activity while the matrix and membrane insoluble protein did not displayed AOX activity. The proteins were purified with FPLC by adding deoxycholamide dBC. The purified enzyme fraction exhibited a high specific activity, which could be kept for at least 6 months when stored at -70 degrees C. Furthermore, we used a silver stain system to identify the AOX, which showed 4 different protein bands from 30 kD to 32 kD. 2-dimensional electrophoresis showed 4 isoelectric points in the range from pH6.4 to 7.4 respectively.
Insights
This study found abundant alternate oxidase (AOX) in thermogenic plants. Purified AOX protein fractions showed high activity and stability, with identified protein bands and isoelectric points.
Area of Science:
- Biochemistry
- Plant Physiology
Context:
- Thermogenic plants, such as *Arum maculatum*, possess abundant alternate oxidase (AOX).
- Mitochondria isolated from *Arum maculatum* inflorescences exhibit high oxygen consumption rates.
Purpose:
- To isolate, purify, and characterize the alternate oxidase (AOX) enzyme from thermogenic *Arum maculatum*.
- To determine the location and properties of AOX within isolated mitochondria.
Summary:
- Mitochondria isolated from *Arum maculatum* inflorescences demonstrated significant oxygen consumption. AOX activity was localized to the mitochondrial membrane and soluble membrane proteins, but not the matrix or insoluble membrane proteins.
- Purification via FPLC yielded a highly active AOX enzyme fraction, stable for at least six months at -70°C. Silver staining revealed four protein bands between 30-32 kD, and 2D electrophoresis indicated four isoelectric points from pH 6.4 to 7.4.
Impact:
- This research provides a detailed biochemical characterization of AOX in a thermogenic plant.
- The findings contribute to understanding the role of AOX in plant respiration and thermogenesis.
