JBIC Journal of Biological Inorganic Chemistry

, Volume 18, Issue 6, pp 595–598 | Cite as

Building reactive copper centers in human carbonic anhydrase II

  • He Song
  • Andrew C. Weitz
  • Michael P. Hendrich
  • Edwin A. Lewis
  • Joseph P. Emerson


Reengineering metalloproteins to generate new biologically relevant metal centers is an effective a way to test our understanding of the structural and mechanistic features that steer chemical transformations in biological systems. Here, we report thermodynamic data characterizing the formation of two type-2 copper sites in carbonic anhydrase and experimental evidence showing one of these new, copper centers has characteristics similar to a variety of well-characterized copper centers in synthetic models and enzymatic systems. Human carbonic anhydrase II is known to bind two Cu2+ ions; these binding events were explored using modern isothermal titration calorimetry techniques that have become a proven method to accurately measure metal-binding thermodynamic parameters. The two Cu2+-binding events have different affinities (K a approximately 5 × 1012 and 1 × 1010), and both are enthalpically driven processes. Reconstituting these Cu2+ sites under a range of conditions has allowed us to assign the Cu2+-binding event to the three-histidine, native, metal-binding site. Our initial efforts to characterize these Cu2+ sites have yielded data that show distinctive (and noncoupled) EPR signals associated with each copper-binding site and that this reconstituted enzyme can activate hydrogen peroxide to catalyze the oxidation of 2-aminophenol.


Carbonic Anhydrase Isothermal Titration Calorimetry Copper Center Dipicolinic Acid Dicopper 
These keywords were added by machine and not by the authors. This process is experimental and the keywords may be updated as the learning algorithm improves.

Supplementary material

775_2013_1009_MOESM1_ESM.pdf (241 kb)
Experimental details as well as Figs. A, B, and C can be found in the electronic supplementary material. Supplementary material 1 (PDF 240 kb)


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Copyright information

© SBIC 2013

Authors and Affiliations

  • He Song
    • 1
  • Andrew C. Weitz
    • 2
  • Michael P. Hendrich
    • 2
  • Edwin A. Lewis
    • 1
  • Joseph P. Emerson
    • 1
  1. 1.Department of Chemistry, 1115 Hand LabMississippi State UniversityMississippi StateUSA
  2. 2.Department of ChemistryCarnegie Mellon UniversityPittsburghUSA

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