FerB is a flavoenzyme capable of reducing quinones, ferric complexes and chromate. Its expression in Escherichia coli as a hexahistidine fusion resulted in a functional product only when the tag was placed on the C-terminus. The molecular mass values estimated by gel permeation chromatography were compatible with the existence of either dimer or trimer, whereas the light scattering data, together with cross-linking experiments that yielded exclusively monomer and dimer bands on dodecyl sulfate-polyacrylamide gels, strongly supported a dimeric nature of both native and tagged form of FerB. These two proteins also exhibited almost identical secondary structure as judged by Fourier transform infra red spectrometry. The presence of tag, however, shifted the temperature of thermal inactivation as well as the thermal denaturation curve towards lower temperatures. Despite somewhat lower thermal stability, the fusion protein is considered a better candidate for crystallization than the wild-type one due to a more negative value of its second optical viral coefficient.
- MeSH
- diferenciální skenovací kalorimetrie MeSH
- Escherichia coli genetika MeSH
- Fourierova analýza MeSH
- histidin genetika chemie metabolismus MeSH
- multimerizace proteinu MeSH
- NADH, NADPH oxidoreduktasy biosyntéza genetika chemie metabolismus MeSH
- NADP metabolismus MeSH
- oligopeptidy genetika chemie metabolismus MeSH
- Paracoccus denitrificans enzymologie genetika MeSH
- rekombinantní fúzní proteiny biosyntéza genetika chemie metabolismus MeSH
- sekundární struktura proteinů MeSH
- stabilita enzymů MeSH
- teplota MeSH
- Publikační typ
- práce podpořená grantem MeSH
