Related Experiment Video
Updated: Jun 11, 2026

Application of Genetically Encoded Fluorescent Nitric Oxide (NO•) Probes, the geNOps, for Real-time Imaging of NO• Signals in Single Cells
Published on: March 16, 2017
Introducing a 2-His-1-Glu nonheme iron center into myoglobin confers nitric oxide reductase activity
Ying-Wu Lin1, Natasha Yeung, Yi-Gui Gao
1Department of Chemistry, University of Illinois at Urbana-Champaign, Urbana, Illinois 61801, USA.
Abstract:
A conserved 2-His-1-Glu metal center, as found in natural nonheme iron-containing enzymes, was engineered into sperm whale myoglobin by replacing Leu29 and Phe43 with Glu and His, respectively (swMb L29E, F43H, H64, called Fe(B)Mb(-His)). A high resolution (1.65 A) crystal structure of Cu(II)-CN(-)-Fe(B)Mb(-His) was determined, demonstrating that the unique 2-His-1-Glu metal center was successfully created within swMb. The Fe(B)Mb(-His) can bind Cu, Fe, or Zn ions, with both Cu(I)-Fe(B)Mb(-His) and Fe(II)-Fe(B)Mb(-His) exhibiting nitric oxide reductase (NOR) activities. Cu dependent NOR activity was significantly higher than that of Fe in the same metal binding site. EPR studies showed that the reduction of NO to N(2)O catalyzed by these two enzymes resulted in different intermediates; a five-coordinate heme-NO species was observed for Cu(I)-Fe(B)Mb(-His) due to the cleavage of the proximal heme Fe-His bond, while Fe(II)-Fe(B)Mb(-His) remained six-coordinate. Therefore, both the metal ligand, Glu29, and the metal itself, Cu or Fe, play crucial roles in NOR activity. This study presents a novel protein model of NOR and provides insights into a newly discovered member of the NOR family, gNOR.
More Related Videos
08:23Chemiluminescence-based Assays for Detection of Nitric Oxide and its Derivatives from Autoxidation and Nitrosated Compounds
Published on: February 16, 2022
08:02Benchtop Immobilized Metal Affinity Chromatography, Reconstitution and Assay of a Polyhistidine Tagged Metalloenzyme for the Undergraduate Laboratory
Published on: August 23, 2018
Related Concept Videos
Hemoglobin
When all four heme groups are bound to oxygen, the resulting molecule is called oxyhemoglobin. As a result, arterial blood...
Nitric Oxide Signaling Pathway
Electron Transport Chain: Complex I and II
ROS generation is regulated and maintained at moderate levels necessary...
Protein and Protein Structure
A protein's shape is critical to its function. For example, an enzyme can...
Inorganic Nitrogen Assimilation
Gene Families
Occasionally these regions can be adapted to take on new roles within the organism, becoming novel genes...