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A spectroscopic characterization of a monomeric analog of copper, zinc superoxide dismutase

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Abstract

A mutated protein of human Cu(II)2Zn(II)2 SOD in which residues Phe50 and Gly51 at the dimer interface were substituted by Glu's, thus producing a monomeric species, has been characterized by electronic absorption spectroscopy, EPR, relaxivity and1H NMR techniques. Such substitutions and/or accompanying remodeling and exposure of the dimer interface to solvent, alter the geometry of the active site: increases in the axiality of the copper chromophore and the Cu-OH2 distance have been observed. The affinity of both metal binding sites for Co(II) is also altered. The observed NMR parameters of the Co(II) substituted derivative have been interpreted as a function of the decrease of rotational correlation time as a consequence of the lower molecular weight of the mutated protein. Sharper NMR signals are also obtained for the reduced diamagnetic enzyme. Results are consistent with an active site structure similar to that observed for the dimeric analog Thr137Ile characterized elsewhere. An observed proportional decrease in enzymatic activity and affinity for the N3-anion suggests the importance of electrostatic forces during substrate docking and catalysis.

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Author notes

  1. Guy T. Mullenbach

    Present address: Lynx Therapeutics, 94404, Foster City, CA, USA

Authors and Affiliations

  1. Department of Chemistry, University of Florence, I-50121, Florence, Italy

    Ivano Bertini, Mario Piccioli & Maria Silvia Viezzoli

  2. Chiron Corporation, 94608, Emeryville, CA, USA

    Choi Ying Chiu & Guy T. Mullenbach

Authors
  1. Ivano Bertini
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  2. Mario Piccioli
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  3. Maria Silvia Viezzoli
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  4. Choi Ying Chiu
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  5. Guy T. Mullenbach
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Bertini, I., Piccioli, M., Viezzoli, M.S. et al. A spectroscopic characterization of a monomeric analog of copper, zinc superoxide dismutase. Eur Biophys J 23, 167–176 (1994). https://doi.org/10.1007/BF01007608

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  • DOI: https://doi.org/10.1007/BF01007608

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