Welcome to the new version of CaltechAUTHORS. Login is currently restricted to library staff. If you notice any issues, please email coda@library.caltech.edu
Published August 2018 | Supplemental Material
Journal Article Open

Functional importance of Glutamate-445 and Glutamate-99 in proton-coupled electron transfer during oxygen reduction by cytochrome bd from Escherichia coli

Abstract

The recent X-ray structure of the cytochrome bd respiratory oxygen reductase showed that two of the three heme components, heme d and heme b, have glutamic acid as an axial ligand. No other native heme proteins are known to have glutamic acid axial ligands. In this work, site-directed mutagenesis is used to probe the roles of these glutamic acids, E445 and E99 in the E. coli enzyme. It is concluded that neither glutamate is a strong ligand to the heme Fe and they are not the major determinates of heme binding to the protein. Although very important, neither glutamate is absolutely essential for catalytic function. The close interactions between the three hemes in cyt bd result in highly cooperative properties. For example, mutation of E445, which is near heme d, has its greatest effects on the properties of heme b and heme b. It is concluded that 1) O binds to the hydrophilic side of heme d and displaces E445; 2) E445 forms a salt bridge with R448 within the O binding pocket, and both residues play a role to stabilize oxygenated states of heme d during catalysis; 3) E445 and E99 are each protonated accompanying electron transfer to heme d and heme b, respectively; 4) All protons used to generate water within the heme d active site come from the cytoplasm and are delivered through a channel that must include internal water molecules to assist proton transfer: [cytoplasm] → E107 → E99 (heme b) → E445 (heme d) → oxygenated heme d.

Additional Information

© 2018 Elsevier. Received 12 February 2018, Revised 25 April 2018, Accepted 26 April 2018, Available online 30 April 2018.

Attached Files

Supplemental Material - 1-s2.0-S0005272818300884-mmc1.pdf

Supplemental Material - 1-s2.0-S0005272818300884-mmc2.docx

Files

1-s2.0-S0005272818300884-mmc1.pdf
Files (2.9 MB)
Name Size Download all
md5:8fa8c5e3d5a34c5deaee0a8404319058
544.7 kB Download
md5:16acd52a284c2d5e5392832de337dbe9
2.4 MB Preview Download

Additional details

Created:
August 21, 2023
Modified:
October 18, 2023