期刊
JOURNAL OF INORGANIC BIOCHEMISTRY
卷 162, 期 -, 页码 164-177出版社
ELSEVIER SCIENCE INC
DOI: 10.1016/j.jinorgbio.2016.04.019
关键词
Ferritin; Di-metal carboxylate protein; Ribonucleotide reductase; EPR spectroscopy; Mossbauer spectroscopy; X-ray crystallography
资金
- Max Planck Gesellschaft
- Cluster of Excellence RESOLV - Deutsche Forschungsgemeinschaft [EXC 1069]
- Australian Research Council [FT140100834]
- Alexander von Humboldt Foundation
- Deutsche Forschungsgemeinschaft [Ha3265/6-1]
- Deutsche Forschungsgemeinschaft (Heisenberg Fellowship)
- German Bundesministerium fur Bildung and Forschung within the Rontgen-Angstrom Cluster [05K14KE1]
- Swedish Research Council
- Swedish Foundation for Strategic Research
- Swedish Cancer Society
- Knut and Alice Wallenberg Foundation
- European Community [283570]
A manganese/iron cofactor which performs multi-electron oxidative chemistry is found in two classes of ferritin-like proteins, the small subunit (R2) of dass Ic ribonucleotide reductase (R2c) and the R2-like ligand-binding oxidase (R2lox). It is undear how a heterodimeric Mn/Fe metallocofactor is assembled in these two related proteins as opposed to a homodimeric Fe/Fe cofactor, especially considering the structural similarity and proximity of the two metal-binding sites in both protein scaffolds and the similar first coordination sphere ligand preferences of Mn-II and Fe-II. Using EPR and Mfissbauer spectroscopies as well as X-ray anomalous dispersion, we examined metal loading and cofactor activation of both proteins in vitro (in solution). We find divergent cofactor assembly mechanisms for the two systems. In both cases, excess Mn-II promotes heterobimetallic cofactor assembly. In the absence of Fe-II, R2c cooperatively binds Mn-II at both metal sites, whereas R2lox does not readily bind Mn-II at either site. Heterometallic cofactor assembly is favored at substoichiometric Feu concentrations in R2lox. Fe-II and Mn-II likely bind to the protein in a stepwise fashion, with Feu binding to site 2 initiating cofactor assembly. In R2c, however, heterometallic assembly is presumably achieved by the displacement of Mn-II by Fe-II at site 2. The divergent metal loading mechanisms are correlated with the putative in vivo functions of R2c and R2lox, and most likely with the intracellular Mn-II/Fe-II concentrations in the host organisms from which they were isolated. (C) 2016 Elsevier Inc. All rights reserved.
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