6F6M

R2-like ligand-binding oxidase Y162F mutant with anaerobically reconstituted Mn/Fe cofactor

Summary for 6F6M

• Entry DOI: 10.2210/pdb6f6m/pdb
• Related: 4HR0 4HR4 4HR5 4XB9 4XBV 4XBW 5DCO 5DCR 5DCS 5EKB 5OMJ 5OMK 6F65 6F6B 6F6C 6F6E 6F6F 6F6G 6F6H 6F6K 6F6L
• Descriptor: Ribonucleotide reductase small subunit, FE (II) ION, MANGANESE (II) ION, ... (5 entities in total)
• Functional Keywords: r2-like ligand-binding oxidase, mn/fe cofactor, ribonucleotide reductase r2 subunit fold, metalloprotein oxidoreductase, oxidoreductase
• Biological source: Geobacillus kaustophilus (strain HTA426)
• Total number of polymer chains: 2
• Total formula weight: 74739.72

Authors

Griese, J.J.,Hogbom, M. (deposition date: 2017-12-05, release date: 2018-12-19, Last modification date: 2024-01-17)

Primary citation

Griese, J.J.,Kositzki, R.,Haumann, M.,Hogbom, M.
Assembly of a heterodinuclear Mn/Fe cofactor is coupled to tyrosine-valine ether cross-link formation in the R2-like ligand-binding oxidase.
J. Biol. Inorg. Chem., 24:211-221, 2019

PubMed Abstract: R2-like ligand-binding oxidases (R2lox) assemble a heterodinuclear Mn/Fe cofactor which performs reductive dioxygen (O) activation, catalyzes formation of a tyrosine-valine ether cross-link in the protein scaffold, and binds a fatty acid in a putative substrate channel. We have previously shown that the N-terminal metal binding site 1 is unspecific for manganese or iron in the absence of O, but prefers manganese in the presence of O, whereas the C-terminal site 2 is specific for iron. Here, we analyze the effects of amino acid exchanges in the cofactor environment on cofactor assembly and metalation specificity using X-ray crystallography, X-ray absorption spectroscopy, and metal quantification. We find that exchange of either the cross-linking tyrosine or the valine, regardless of whether the mutation still allows cross-link formation or not, results in unspecific manganese or iron binding at site 1 both in the absence or presence of O, while site 2 still prefers iron as in the wild-type. In contrast, a mutation that blocks binding of the fatty acid does not affect the metal specificity of either site under anoxic or aerobic conditions, and cross-link formation is still observed. All variants assemble a dinuclear trivalent metal cofactor in the aerobic resting state, independently of cross-link formation. These findings imply that the cross-link residues are required to achieve the preference for manganese in site 1 in the presence of O. The metalation specificity, therefore, appears to be established during the redox reactions leading to cross-link formation.

PubMed: 30689052
DOI: 10.1007/s00775-019-01639-4

Experimental method

X-RAY DIFFRACTION (1.393 Å)