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The Multifunctional Catalytic Hemoglobin from Amphitrite ornata: Protocols on Isolation, Taxonomic Identification,
Anna L Husted1, Victoria R Sutton1, Lauren A Presnar1
1Department of Natural Sciences, University of South Carolina Beaufort, 1 University Boulevard, Bluffton, SC 29909, USA.
Methods and Protocols
|December 27, 2024
Summary
Researchers developed a DNA barcoding method to easily identify the polychaete worm Amphitrite ornata. This facilitates finding new dehaloperoxidases (AoDHP) with unique catalytic abilities in marine invertebrates.
Area of Science:
- Biochemistry
- Marine Biology
- Genetics
Background:
- The terebellid polychaete Amphitrite ornata possesses a multifunctional dehaloperoxidase (AoDHP) with oxygen transport and substrate oxidation activities.
- Identifying novel dehaloperoxidases from macrobenthic infauna is challenging due to taxonomic identification difficulties.
Purpose of the Study:
- To develop a simplified method for accurate taxonomic identification of Amphitrite ornata.
- To facilitate the discovery of new dehaloperoxidase candidates in marine invertebrates.
Main Methods:
- DNA barcoding using the mitochondrial cytochrome c oxidase subunit I gene for Amphitrite ornata identification.
- Extraction and purification of AoDHP from whole worm specimens.
- Overexpression of recombinant AoDHP isoenzyme A in E. coli for activity comparison.
Main Results:
- A streamlined DNA barcoding method for Amphitrite ornata identification was established, suitable for non-specialists.
- Peroxidase activity assays showed a strong correlation between native and recombinant AoDHP Michaelis-Menten parameters.
- The study successfully purified and characterized AoDHP from Amphitrite ornata.
Conclusions:
- The described method simplifies the taxonomic identification of Amphitrite ornata, aiding dehaloperoxidase discovery.
- This approach is predicted to accelerate the identification of novel dehaloperoxidases from other marine invertebrates.
Keywords:
DNA barcodingUV absorbance-based assayallelochemicaldehaloperoxidasehalogenated aromatic compoundhemoglobininfaunamarine benthic ecologymolecular taxonomypolychaeteMore Related Videos
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