Abstract
THE enzyme Cu, Zn superoxide dismutase (SOD) protects against oxidative damage by dismuting the superoxide radical O.−2 to molecular oxygen and hydrogen peroxide1–3 at the active-site Cuion4,5 in a reaction that is rate-limited by diffusion3,6 and enhanced by electrostatic guidance7–10. SOD has evolved to be one of the fastest enzymes known ( Vmax ~ 2 x 109 M−1 s−1)6,11. The new crystal structures of human SOD12 show that amino-acid site chains that are implicated in electrostatic guidance8 (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→Gin) 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→Gin) 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.
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Getzoff, E., Cabelli, D., Fisher, C. et al. Faster superoxide dismutase mutants designed by enhancing electrostatic guidance. Nature 358, 347–351 (1992). https://doi.org/10.1038/358347a0
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DOI: https://doi.org/10.1038/358347a0
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