X-ray Spectroscopy of Nitrile Hydratase at pH 7 and 9
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- 作者：Robert C. Scarrow ; Bridget A. Brennan ; John G. Cummings ; Haiyong Jin ; Daihung J. Duong ; James T. Kindt ; and Mark J. Nelson
- 刊名：Biochemistry
- 出版年：1996
- 出版时间：August 6, 1996
- 年：1996
- 卷：35
- 期：31
- 页码：10078 - 10088
- 全文大小：461K
- 年卷期：v.35,no.31(August 6, 1996)
- ISSN：1520-4995
文摘
The iron K-edge X-ray absorption spectrum ofRhodococcus sp. R312 (formerlyBrevibacteriumsp. R312) nitrile hydratase in frozen solutions at pH 7 and 9 has beenanalyzed to determine details of theiron coordination. EXAFS analysis implies two or three sulfurligands per iron and overall six coordination;together with previous EPR and ENDOR results, this implies anN3S2O ligation sphere. The bondlengthsfrom EXAFS analysis [rav(Fe-S) = 2.21Å at pH 7.3; rav(Fe-N/O) = 1.99 Å]support cis coordinationof two cysteine ligands and conclusively rule out nitric oxidecoordination to the iron, a possibility proposedon the basis of an FTIR difference experiment [Noguchi, T., Honda, J.,Nagamune, T., Sasabe, H., Inoue,Y., & Endo, I. (1995) FEBS Lett. 358,9-12]. The higher-frequency EXAFS can be simulated wellbyinclusion of multiple scattering from two or three imidazole ligands;the fit to the data is improved iffirst-sphere multiple scattering pathways are also included. Aslight shortening (by 0.02 ± 0.01 Å) ofone or both Fe-S bonds when the pH is raised from 7.3 to 9.0 isconsistent with shifts observed in theRaman spectrum [Brennan et al. (1996) Biochemistry35, 10068-10077].