Stability of spermine oxidase to thermal and chemical denaturation: comparison with bovine serum amine oxidase
| Autor: | Laura Cervoni, Shinji Ohkubo, Rodolfo Federico, Alessia Leonetti, Marla Xhani, Pasquale Stano, Enzo Agostinelli, Fabio Polticelli, Paolo Mariottini, Manuela Cervelli |
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| Přispěvatelé: | Cervelli, Manuela, Leonetti, Alessia, Cervoni, Laura, Ohkubo, Shinji, Xhani, Marla, Stano, Pasquale, Federico, Rodolfo, Polticelli, Fabio, Mariottini, Paolo, Agostinelli, Enzo |
| Rok vydání: | 2016 |
| Předmět: |
0301 basic medicine
Protein Denaturation Amine oxidase Spermine oxidase Clinical Biochemistry Spermine Biochemistry 03 medical and health sciences chemistry.chemical_compound Enzyme Stability Animals Quaternary structure Denaturation (biochemistry) Bovine serum albumin Enzyme stability Oxidoreductases Acting on CH-NH Group Donors biology Organic Chemistry Bovine serum amine oxidase Amine oxidase (copper-containing) Recombinant Proteins 030104 developmental biology chemistry biology.protein Cattle Protein quaternary structure Amine Oxidase (Copper-Containing) Protein Multimerization Polyamine oxidase |
| Zdroj: | Amino Acids. 48:2283-2291 |
| ISSN: | 1438-2199 0939-4451 |
| DOI: | 10.1007/s00726-016-2273-5 |
| Popis: | Spermine oxidase (SMOX) is a flavin-containing enzyme that specifically oxidizes spermine to produce spermidine, 3-aminopropanaldehyde and hydrogen peroxide. While no crystal structure is available for any mammalian SMOX, X-ray crystallography showed that the yeast Fms1 polyamine oxidase has a dimeric structure. Based on this scenario, we have investigated the quaternary structure of the SMOX protein by native gel electrophoresis, which revealed a composite gel band pattern, suggesting the formation of protein complexes. All high-order protein complexes are sensitive to reducing conditions, showing that disulfide bonds were responsible for protein complexes formation. The major gel band other than the SMOX monomer is the covalent SMOX homodimer, which was disassembled by increasing the reducing conditions, while being resistant to other denaturing conditions. Homodimeric and monomeric SMOXs are catalytically active, as revealed after gel staining for enzymatic activity. An engineered SMOX mutant deprived of all but two cysteine residues was prepared and characterized experimentally, resulting in a monomeric species. High-sensitivity differential scanning calorimetry of SMOX was compared with that of bovine serum amine oxidase, to analyse their thermal stability. Furthermore, enzymatic activity assays and fluorescence spectroscopy were used to gain insight into the unfolding process. |
| Databáze: | OpenAIRE |
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