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Faster superoxide-dismutase mutants designed by enhancing electrostatic guidance

Academic Article
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Overview

authors

  • Getzoff, Elizabeth
  • Cabelli, D. E.
  • Fisher, C. L.
  • Parge, H. E.
  • Viezzoli, M. S.
  • Banci, L.
  • Hallewell, R. A.

publication date

  • 1992

journal

  • Nature  Journal

abstract

  • The enzyme Cu, Zn superoxide dismutase (SOD) protects against oxidative damage by dismuting the superoxide radical O2-. to molecular oxygen and hydrogen peroxide at the active-site Cu ion in a reaction that is rate-limited by diffusion and enhanced by electrostatic guidance. SOD has evolved to be one of the fastest enzymes known (V(max) approximately 2 x 10(9) M-1 s-1). The new crystal structures of human SOD show that amino-acid site chains that are implicated in electrostatic guidance (Glu 132, Glu 133 and Lys 136) form a hydrogen-bonding network. Here we show that site-specific mutants that increase local positive charge while maintaining this orienting network (Glu----Gln) have faster reaction rates and increased ionic-strength dependence, matching brownian dynamics simulations incorporating electrostatic terms. Increased positive charge alone is insufficient: one charge reversal (Glu----Lys) mutant is slower than the equivalent charge neutralization (Glu----Gln) mutant, showing that the newly introduced positive charge disrupts the orienting network. Thus, electrostatically facilitated diffusion rates can be increased by design, provided the detailed structural integrity of the active-site electrostatic network is maintained.

subject areas

  • Amino Acid Sequence
  • Computer Simulation
  • Electrochemistry
  • Glutamates
  • Glutamic Acid
  • Humans
  • Hydrogen Bonding
  • Hydrogen-Ion Concentration
  • Kinetics
  • Lysine
  • Models, Molecular
  • Mutagenesis, Site-Directed
  • Osmolar Concentration
  • Protein Conformation
  • Superoxide Dismutase
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Identity

International Standard Serial Number (ISSN)

  • 0028-0836

Digital Object Identifier (DOI)

  • 10.1038/358347a0

PubMed ID

  • 1353610
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Additional Document Info

start page

  • 347

end page

  • 351

volume

  • 358

issue

  • 6384

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