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Partial purification of L-ascorbate:ferricytochrome b5 oxidoreductase from rat liver microsomes
Hoppe-Seyler'S Zeitschrift Fur Physiologische Chemie
|November 1, 1977
Abstract:
L-Ascorbate:ferricytochrome b5 oxidoreductase from rat liver microsomes was enriched about 750-fold after treatment with a non-ionic detergent with a yield of about 15%. The enriched enzyme still shows four bands in sodium dodecylsulfate polyacrylamide-gel electrophoresis and is a hydrophobic membrane protein.
Insights
Researchers enriched L-Ascorbate:ferricytochrome b5 oxidoreductase from rat liver microsomes using a non-ionic detergent. The hydrophobic membrane protein was purified 750-fold, retaining its activity.
Area of Science:
- Biochemistry
- Enzymology
- Membrane Protein Research
Background:
- Microsomal fractions are crucial for cellular redox reactions.
- L-Ascorbate:ferricytochrome b5 oxidoreductase plays a role in electron transport.
- Hydrophobic membrane proteins are challenging to isolate and study.
Purpose of the Study:
- To enrich and characterize L-Ascorbate:ferricytochrome b5 oxidoreductase from rat liver microsomes.
- To investigate the properties of the purified enzyme.
- To confirm the enzyme's hydrophobic nature.
Main Methods:
- Enzymatic enrichment using non-ionic detergents.
- Sodium dodecylsulfate polyacrylamide-gel electrophoresis (SDS-PAGE) for purity assessment.
- Characterization of enzyme properties.
Main Results:
- Achieved a 750-fold enrichment of the enzyme.
- Obtained a yield of approximately 15% for the purified enzyme.
- SDS-PAGE revealed four distinct protein bands in the enriched fraction.
- Confirmed the enzyme as a hydrophobic membrane protein.
Conclusions:
- Successful enrichment of L-Ascorbate:ferricytochrome b5 oxidoreductase was achieved.
- The purified enzyme exhibits multiple polypeptide chains and is intrinsically hydrophobic.
- This provides a foundation for further functional and structural studies of this enzyme.