Rapid-freeze-quench magnetic circular dichroism of intermediate X in ribonucleotide reductase: new structural insight.
To elucidate the electronic structure of intermediate X in the oxygen activation reaction of the R2 subunit of ribonucleotide reductase, a protocol has been developed to perform magnetic circular dichroism (MCD) on a rapid-freeze-quench, strain free optical sample. RFQ-MCD data have been collected on intermediate X in the double mutant of R2, Y122/Y356F. While X has been reported to exhibit a broad absorption band at 365 nm, there are at least 10 electronic transitions observed at low-temperature MCD. From C0/D0 ratios, the transitions of X can be divided into three regions: 16 000-22 000 cm-1 region involving spin-allowed ligand field transitions of the Fe(IV), 23 000-24 000 cm-1 region of spin-forbidden, spin-flip transitions on the Fe(IV), and the charge transfer (CT) region from 26 000 to 32 000 cm-1. The C0/D0 ratios from d --> d and CT transitions strongly support significant Fe(IV) character coupled into the paramagnetic center. Ligand field (spin-allowed d --> d region) analysis allows the bis-mu-oxo and mu-oxo plus other monoanionic bridge possibilities for the structure of intermediate X to be distinguished, providing new insight into the molecular mechanism of the cluster formation in R2.
Mitić
Natasa
N
Department of Chemistry, Stanford University, 333 Campus Street, Stanford, California 94305, USA.
Saleh
Lana
L
Schenk
Gerhard
G
Bollinger
J Martin
JM
Solomon
Edward I
EI
D002942:Circular Dichroism / Q000379:methods*; D004735:Energy Transfer; D005615:Freezing; D007501:Iron / Q000737:chemistry; D012264:Ribonucleotide Reductases / Q000737:chemistry* / Q000378:metabolism*; D013997:Time Factors
D007501:Iron; D012264:Ribonucleotide Reductases