Modulation of the Redox Potential of the [Fe(SCys)4] Site in Rubredoxin by the Orientation of a Peptide Dipole
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- 作者：Marly K. Eidsness ; Amy E. Burden ; Kimberly A. Richie ; Donald M. Kurtz ; Jr. ; Robert A. Scott ; Eugene T. Smith ; Toshiko Ichiye ; Brian Beard ; TongPil Min ; and ChulHee Kang
- 刊名：Biochemistry
- 出版年：1999
- 出版时间：November 9, 1999
- 年：1999
- 卷：38
- 期：45
- 页码：14803 - 14809
- 全文大小：184K
- 年卷期：v.38,no.45(November 9, 1999)
- ISSN：1520-4995
文摘
Rubredoxins (Rds) may be separated into two classes based upon the correlation of theirreduction potentials with the identity of residue 44; those with Ala44 have reduction potentials that are~50 mV higher than those with Val44. The smaller side chain volume occupied by Ala44 relative to thatoccupied by Val44 has been proposed to explain the increase in the reduction potential, based upon changesin the Gly43-Ala44 peptide bond orientation and the distance to the [Fe(SCys)4] center in the Pyrococcusfuriosus (Pf) Rd crystal structure compared to those of Gly43-Val44 in the Clostridium pasteurianum(Cp) Rd crystal structure. As an experimental test of this hypothesis, single-site Val44 
Ala44 exchangemutants, [V44A]Cp and [A44V]Pf Rds, have been cloned and expressed. Reduction potentials of theseresidue 44 variants and pertinent features of the X-ray crystal structure of [V44A]Cp Rd are reported.Relative to those of wild-type Cp and Pf Rds, the V44A mutation in Cp Rd results in an 86 mV increasein midpoint reduction potential and the [A44V] mutation in Pf Rd results in a 95 mV decrease in midpointreduction potential, respectively. In the crystal structure of [V44A]Cp Rd, the peptide bond between residues43 and 44 is ~0.3 Å closer to the Fe center and the hydrogen bond distance between the residue 44peptide nitrogen and the Cys42 
-sulfur decreases by 0.32 Å compared to the analogous distances in thewild-type Cp Rd crystal structure. The results described herein support the prediction that the identity ofresidue 44 alone determines whether a Rd reduction potential of about -50 or 0 mV is observed.
Ala44 exchangemutants, [V44A]Cp and [A44V]Pf Rds, have been cloned and expressed. Reduction potentials of theseresidue 44 variants and pertinent features of the X-ray crystal structure of [V44A]Cp Rd are reported.Relative to those of wild-type Cp and Pf Rds, the V44A mutation in Cp Rd results in an 86 mV increasein midpoint reduction potential and the [A44V] mutation in Pf Rd results in a 95 mV decrease in midpointreduction potential, respectively. In the crystal structure of [V44A]Cp Rd, the peptide bond between residues43 and 44 is ~0.3 Å closer to the Fe center and the hydrogen bond distance between the residue 44peptide nitrogen and the Cys42 -sulfur decreases by 0.32 Å compared to the analogous distances in thewild-type Cp Rd crystal structure. The results described herein support the prediction that the identity ofresidue 44 alone determines whether a Rd reduction potential of about -50 or 0 mV is observed.