Copper-catalyzed oxidation of the recombinant SHa(29–231) prion protein
Autor: | Stanley B. Prusiner, Earl R. Stadtman, Darlene Groth, Jesús R. Requena, Rodney L. Levine, Giuseppe Legname |
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Rok vydání: | 2001 |
Předmět: |
Prions
Molecular Sequence Data chemistry.chemical_element Ascorbic Acid Oxygen Catalysis Mass Spectrometry law.invention Metal law Settore BIO/10 - Biochimica Humans Amino Acid Sequence Peptide sequence Chromatography High Pressure Liquid Histidine Multidisciplinary Chemistry Biological Sciences Ascorbic acid Copper Peptide Fragments Recombinant Proteins In vitro Kinetics Models Chemical Biochemistry visual_art visual_art.visual_art_medium Recombinant DNA Oxidation-Reduction |
Zdroj: | Proceedings of the National Academy of Sciences. 98:7170-7175 |
ISSN: | 1091-6490 0027-8424 |
Popis: | Metal-catalyzed oxidation may result in structural damage to proteins and has been implicated in aging and disease, including neurological disorders such as Alzheimer's disease and amyotrophic lateral sclerosis. The selective modification of specific amino acid residues with high metal ion affinity leads to subtle structural changes that are not easy to detect but may have dramatic consequences on physical and functional properties of the oxidized protein molecules. PrP contains a histidine-rich octarepeat domain that binds copper. Because copper-binding histidine residues are particularly prone to metal-catalyzed oxidation, we investigated the effect of this reaction on the recombinant prion protein SHaPrP(29–231). Using Cu 2+ /ascorbate, we oxidized SHaPrP(29–231) in vitro . Oxidation was demonstrated by liquid chromatography/mass spectrometry, which showed the appearance of protein species of higher mass, including increases in multiples of 16, characteristic of oxygen incorporation. Digestion studies using Lys C indicate that the 29–101 region, which includes the histidine-containing octarepeats, is particularly affected by oxidation. Oxidation was time- and copper concentration-dependent and was evident with copper concentrations as low as 1 μM. Concomitant with oxidation, SHaPrP(29–231) suffered aggregation and precipitation, which was nearly complete after 15 min, when the prion protein was incubated at 37°C with a 6-fold molar excess of Cu 2+ . These findings indicate that PrP, a copper-binding protein, may be particularly susceptible to metal-catalyzed oxidation and that oxidation triggers an extensive structural transition leading to aggregation. |
Databáze: | OpenAIRE |
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