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X-Ray Crystallography of Carbon Monoxide Dehydrogenases

  • Jae-Hun Jeoung
  • Berta M. Martins
  • Holger Dobbek
Protocol
Part of the Methods in Molecular Biology book series (MIMB, volume 1876)

Abstract

Carbon monoxide dehydrogenases (CODHs) are central players in the biogeochemical carbon monoxide (CO) cycle and have been extensively studied from the ecological level to the structural/molecular level. Of the two types of CODHs, the oxygen-tolerant CODHs use a bimetallic [CuSMo(=O)OH] center connected to the protein via a pyranopterin cofactor, whereas the oxygen-sensitive CODHs contain a [NiFe4S4-OHx]-cluster. Despite the fact that we have a basic understanding of how both types of CODHs use distinct active sites to catalyze the oxidation of CO with water to CO2, two protons, and two electrons (a reversible reaction in the cases of the oxygen-sensitive CODHs), many questions remain unanswered, especially concerning the electronic structures of the intermediate states. Since these states will likely be only revealed by the interplay of experimental and theoretical methods, there is a need to obtain accurate descriptions of the active site architectures in various states and, consequently, a need to generate crystals with good diffraction quality and collect data at element-specific wavelengths in order to determine the identity of elements in the case of mixed states. This chapter provides a description of the general working protocols for the crystallization and structural analysis of Cu,Mo-CODH and Ni,Fe-CODH that facilitates the mechanistic investigations of these important metalloenzymes.

Key words

Carbon monoxide dehydrogenase CODH Reaction intermediate Transient states Intermediate trapping Crystal dehydration 

Notes

Acknowledgments

The authors are supported by the German funding agency DFG project grants (DO-785/1, DO-787/5, DO-785/6) and the Cluster of Excellence “Unifying Concepts in Catalysis – UniCat” (EXC 314).

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Copyright information

© Springer Science+Business Media, LLC, part of Springer Nature 2019

Authors and Affiliations

  • Jae-Hun Jeoung
    • 1
  • Berta M. Martins
    • 1
  • Holger Dobbek
    • 1
  1. 1.Institute of Biology, Structural Biology and BiochemistryHumboldt-Universität zu BerlinBerlinGermany

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