Design of Heteronuclear Metalloenzymes

A. Bhagi-Damodaran, P. Hosseinzadeh, E. Mirts, J. Reed, I. D. Petrik, Y. Lu

Research output: Chapter in Book/Report/Conference proceedingChapter

7 Scopus citations


Heteronuclear metalloenzymes catalyze some of the most fundamentally interesting and practically useful reactions in nature. However, the presence of two or more metal ions in close proximity in these enzymes makes them more difficult to prepare and study than homonuclear metalloenzymes. To meet these challenges, heteronuclear metal centers have been designed into small and stable proteins with rigid scaffolds to understand how these heteronuclear centers are constructed and the mechanism of their function. This chapter describes methods for designing heterobinuclear metal centers in a protein scaffold by giving specific examples of a few heme–nonheme bimetallic centers engineered in myoglobin and cytochrome c peroxidase. We provide step-by-step procedures on how to choose the protein scaffold, design a heterobinuclear metal center in the protein scaffold computationally, incorporate metal ions into the protein, and characterize the resulting metalloproteins, both structurally and functionally. Finally, we discuss how an initial design can be further improved by rationally tuning its secondary coordination sphere, electron/proton transfer rates, and the substrate affinity.

Original languageEnglish (US)
Title of host publicationMethods in Enzymology
PublisherAcademic Press Inc.
Number of pages37
StatePublished - 2016
Externally publishedYes

Publication series

NameMethods in Enzymology
ISSN (Print)0076-6879
ISSN (Electronic)1557-7988

Bibliographical note

Publisher Copyright:
© 2016 Elsevier Inc.


  • Biomimetic models
  • Biosynthetic models
  • Heme–copper oxidase
  • Manganese peroxidase
  • Metalloprotein design
  • Nitric oxide reductase
  • Oxygen activation
  • Protein design
  • Protein engineering
  • Secondary sphere interactions


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