Proline dehydrogenase from Thermus thermophilus does not discriminate between FAD and FMN as cofactor
| Autor: | Marta Martínez-Júlvez, Adrie H. Westphal, Milagros Medina, Mieke M.E. Huijbers, Willem J. H. van Berkel, Estela Delgado-Arciniega |
|---|---|
| Rok vydání: | 2016 |
| Předmět: |
0301 basic medicine
Models Molecular Rossmann fold animal structures Flavin Mononucleotide Protein Conformation Biochemie Riboflavin Flavin group Crystallography X-Ray Biochemistry Cofactor Maltose-Binding Proteins Article 03 medical and health sciences Proline dehydrogenase Protein structure Bacterial Proteins Oxidoreductase Escherichia coli Proline Oxidase Life Science heterocyclic compounds VLAG chemistry.chemical_classification Multidisciplinary 030102 biochemistry & molecular biology biology Molecular Structure Thermus thermophilus biology.organism_classification Recombinant Proteins enzymes and coenzymes (carbohydrates) Kinetics 030104 developmental biology chemistry biology.protein Flavin-Adenine Dinucleotide bacteria Holoenzymes |
| Zdroj: | Scientific Reports Scientific Reports, 7 Scientific Reports 7 (2017) |
| ISSN: | 2045-2322 |
| Popis: | Flavoenzymes are versatile biocatalysts containing either FAD or FMN as cofactor. FAD often binds to a Rossmann fold, while FMN prefers a TIM-barrel or flavodoxin-like fold. Proline dehydrogenase is denoted as an exception: it possesses a TIM barrel-like fold while binding FAD. Using a riboflavin auxotrophic Escherichia coli strain and maltose-binding protein as solubility tag, we produced the apoprotein of Thermus thermophilus ProDH (MBP-TtProDH). Remarkably, reconstitution with FAD or FMN revealed that MBP-TtProDH has no preference for either of the two prosthetic groups. Kinetic parameters of both holo forms are similar, as are the dissociation constants for FAD and FMN release. Furthermore, we show that the holo form of MBP-TtProDH, as produced in E. coli TOP10 cells, contains about three times more FMN than FAD. In line with this flavin content, the crystal structure of TtProDH variant ΔABC, which lacks helices αA, αB and αC, shows no electron density for an AMP moiety of the cofactor. To the best of our knowledge, this is the first example of a flavoenzyme that does not discriminate between FAD and FMN as cofactor. Therefore, classification of TtProDH as an FAD-binding enzyme should be reconsidered. |
| Databáze: | OpenAIRE |
| Externí odkaz: |
|