Structural and Electronic Factors in Heterolytic Cleavage: Formation of the Co(I) Intermediate in the Corrinoid/Iron-Sulfur Protein from Clostridium thermoaceticum

• Published in: Biochemistry (Easton) 1995-04, Vol.34 (15), p.5269-5273
• Main Authors: Wirt, Michael D, Kumar, Manoj, Wu, Jing-jing, Scheuring, Eva M, Ragsdale, Stephen W, Chance, M. R
• Format: Article
• Language: English
• Subjects: Bacterial Proteins - chemistry; Clostridium - chemistry; Cobalt - chemistry; Iron-Sulfur Proteins - chemistry; Spectrometry, X-Ray Emission; Spectrum Analysis; X-Rays
• ISSN: 0006-2960; 1520-4995
• DOI: 10.1021/bi00015a042

We have completed the first direct structural characterization of an enzyme-bound four-coordinate Co(I) intermediate, in this case for the corrinoid/iron-sulfur protein (C/Fe-SP) from Clostridium thermoaceticum. Extended X-ray absorption fine structure and X-ray edge spectroscopy of the active Co(I) state of the C/Fe-SP indicates a four-coordinate (distorted) square-planar structure where the best fit gives average Co-N(equatorial) distances of 1.87 +/- 0.01 A, corresponding to 4.2 +/- 0.3 ligands. The X-ray edge spectrum of Co(I) C/Fe-SP contains a moderate intensity 1s-4p + "shake-down" (SD) transition and no 1s-3d peak (where SD transitions are indicative of square-planar geometries). X-ray edge results for the methyl-Co(III) form, reported earlier [Wirt, M. D., Kumar, M., Ragsdale, S. W., & Chance, M. R. (1993) J. Am. Chem. Soc. 115, 2146-2150], are consistent with a base-off methylcobamide structure. The absence of a ligated 5-methoxybenzimidazole base in the methyl-Co(III) state is important since the base-off form is predicted to predispose the Co-C bond toward heterolytic cleavage to form the four-coordinate Co(I) species concurrent with methyl transfer. Additionally, we have examined first-derivative X-ray edge spectra of Co(I) C/Fe-SP, relative to edge spectra of a cobalt foil, as an indicator of effective nuclear charge on cobalt. The Co(I) C/Fe-SP edge position at 7720.5 +/- 0.3 eV is less than, but very close to, the value seen for the corresponding free Co(I) cobalamin.