| 1. |
- Andersson, Charlotta S., 1979-, et al.
(author)
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The manganese ion of the heterodinuclear Mn/Fe cofactor in Chlamydia trachomatis ribonucleotide reductase R2c is located at metal position 1.
- 2012
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In: Journal of the American Chemical Society. - : American Chemical Society (ACS). - 0002-7863 .- 1520-5126. ; 134:1, s. 123-125
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Journal article (peer-reviewed)abstract
- The essential catalytic radical of Class-I ribonucleotide reductase is generated and delivered by protein R2, carrying a dinuclear metal cofactor. A new R2 subclass, R2c, prototyped by the Chlamydia trachomatis protein was recently discovered. This protein carries an oxygen-activating heterodinuclear Mn(II)/Fe(II) metal cofactor and generates a radical-equivalent Mn(IV)/Fe(III) oxidation state of the metal site, as opposed to the tyrosyl radical generated by other R2 subclasses. The metal arrangement of the heterodinuclear cofactor remains unknown. Is the metal positioning specific, and if so, where is which ion located? Here we use X-ray crystallography with anomalous scattering to show that the metal arrangement of this cofactor is specific with the manganese ion occupying metal position 1. This is the position proximal to the tyrosyl radical site in other R2 proteins and consistent with the assumption that the high-valent Mn(IV) species functions as a direct substitute for the tyrosyl radical.
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| 2. |
- Leidel, Nils, et al.
(author)
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High-valent [MnFe] and [FeFe] cofactors in ribonucleotide reductases
- 2012
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In: Biochimica et Biophysica Acta - Bioenergetics. - : Elsevier BV. - 0005-2728 .- 1879-2650. ; 1817:3, s. 430-444
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Journal article (peer-reviewed)abstract
- Ribonucleotide reductases (RNRs) are essential for DNA synthesis in most organisms. In class-Ic RNR from Chlamydia trachomatis (Ct), a MnFe cofactor in subunit R2 forms the site required for enzyme activity, instead of an FeFe cofactor plus a redox-active tyrosine in class-la RNRs, for example in mouse (Mus musculus, Mm). For R2 proteins from Ct and Mm, either grown in the presence of, or reconstituted with Mn and Fe ions, structural and electronic properties of higher valence MnFe and FeFe sites were determined by X-ray absorption spectroscopy and complementary techniques, in combination with bond-valence-sum and density functional theory calculations. At least ten different cofactor species could be tentatively distinguished. In Cr R2, two different Mn(IV)Fe(III) site configurations were assigned either L4MnIV(mu O)(2)(FeL4)-L-III (metal-metal distance of similar to 2.75 angstrom, L = ligand) prevailing in metal-grown R2, or L4MnIV(mu O)(mu OH)(FeL4)-L-III (similar to 2.90 angstrom) dominating in metal-reconstituted R2. Specific spectroscopic features were attributed to an Fe(IV)Fe(III) site (similar to 2.55 angstrom) with a L4FeIV(mu O)(2)(FeL3)-L-III core structure. Several Mn,Fe(III)Fe(III) (similar to 2.9-3.1 angstrom) and Mn,Fe(III)Fe(II) species (similar to 3.3-3.4 angstrom) likely showed 5-coordinated Mn(III) or Fe(III). Rapid X-ray photoreduction of iron and shorter metal-metal distances in the high-valent states suggested radiation-induced modifications in most crystal structures of R2. The actual configuration of the MnFe and FeFe cofactors seems to depend on assembly sequences, bound metal type, valence state, and previous catalytic activity involving subunit RI. In Ct R2, the protonation of a bridging oxide in the Mn-IV(mu O)(mu OH)Fe-III core may be important for preventing premature site reduction and initiation of the radical chemistry in R1.
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| 3. |
- Popović-Bijelić, Ana, 1976-
(author)
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Activation and inhibition of diiron and iron-manganese ribonucleotide reductases
- 2012
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Doctoral thesis (other academic/artistic)abstract
- Ribonucleotide reductase (RNR) catalyses the reduction of ribonucleotides to deoxyribonucleotides. In conventional class I RNRs the active site is located in the R1 subunit, and the R2 subunit contains a diiron cofactor and a stable tyrosyl radical essential for activity. Class Ic Chlamydia trachomatis RNR lacks the tyrosyl radical and uses a Mn(IV)Fe(III) cofactor for catalysis. The requirement for metals for RNR activation was studied in C. trachomatis F127Y and Y129F R2, and in mouse wild type and Y177F R2 proteins. The results indicate that R2 affinity for metals is determined by the amino acid located next to the metal site, at the position of the radical carrying tyrosyl. Specifically, R2 proteins that contain phenylalanine in this place bind Mn and Fe, and the tyrosyl containing R2s bind only Fe. Further results show that C. trachomatis RNR can be inhibited by R2 C-terminal oligopeptides. The highest inhibition was observed for a 20-mer peptide indicating that the oligopeptide inhibition mechanism of class Ic is similar to that of the class Ia and b. The second part of the thesis deals with class Ia RNR inhibition. The results show that a lanthanum complex containing three 1,10-phenantroline molecules (KP772) which has showed promising cytotoxic activity in cancer cell lines inhibits mouse R2 protein in the presence of external reductants by iron-chelation. It is suggested that KP772 has several synergistic inhibition mechanisms that contribute to its overall anticancer activity. Moreover, other results show that Triapine, a promising chemotherapeutic compound, and its Fe, Ga, Zn, and Cu complexes, inhibit mouse R2 in both reducing and non-reducing conditions. Inhibition by Triapine involves the binding of the drug to the surface of the R2 protein leading to labilization of the Fe-center and subsequent Fe-chelation by Triapine. Formation of the Fe(II)-Triapine complex which reacts with O2 to produce reactive oxygen species results in complete RNR inactivation.
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