Analysis of protein denaturation, aggregation and post-translational modification by agarose native gel electrophoresis
| Autor: | Tsutomu Arakawa, Yasunori Kurosawa, Chiaki Sakuma, Cynthia Li, Takashi Shibata, Teruo Akuta, Masataka Nakagawa, Yui Tomioka |
|---|---|
| Rok vydání: | 2021 |
| Předmět: |
Protein Denaturation
02 engineering and technology Biochemistry Protein Aggregates Viral Proteins 03 medical and health sciences chemistry.chemical_compound Structural Biology Native state Animals Humans Phosphorylation Bovine serum albumin Molecular Biology 030304 developmental biology Glycated Hemoglobin Gel electrophoresis chemistry.chemical_classification 0303 health sciences ZAP-70 Protein-Tyrosine Kinase biology Transferrin Sodium Dodecyl Sulfate Serum Albumin Bovine Hydrogen Peroxide General Medicine Dependovirus 021001 nanoscience & nanotechnology Blot Electrophoresis chemistry Proteolysis biology.protein Agarose Cattle Electrophoresis Polyacrylamide Gel Hemoglobin 0210 nano-technology Protein Processing Post-Translational |
| Zdroj: | International Journal of Biological Macromolecules. 172:589-596 |
| ISSN: | 0141-8130 |
| Popis: | Agarose native gel electrophoresis has been developed to separate proteins and protein complexes in the native state. Here, we applied this technology to analyze proteins that undergo degradation, post-translational modification or chemical/physical changes. Antibodies showed aggregation/association upon acid or heat treatment. Limited reduction of disulfide bonds resulted in non-covalent aggregation of bovine serum albumin and cleavage of only inter-chain linkages of an antibody that had no effects on its overall structure. Native agarose gel analysis showed changes in mobility of human transferrin upon Fe3+ binding. Analysis of a commercial glycated human hemoglobin A1c showed no difference in electrophoretic pattern from un-modified hemoglobin. Native agarose gel showed aggregation of a virus upon acid or heat treatment. We have extracted bands of bovine serum albumin from the agarose native gel for sodium dodecylsulfate gel electrophoresis analysis, showing degradation of aged sample. Lastly, we analyzed phosphorylation of Zap70 kinase by native gel and Western blotting. These applications should expand the utility of this native gel electrophoresis technology. |
| Databáze: | OpenAIRE |
| Externí odkaz: |
|
Full Text from ScienceDirect