Electron-transfer properties of ruthenated yeast Cu-Zn superoxide dismutase mutants.
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STRUCTURE/FUNCTION 109 B054 ELECTRON-TRANSFER PROPERTIES OF RUTHENATED YEAST CU-ZN SUPEROXIDE DISMUTASE MUTANTS. Yi Lu9h b Jennifer Rennecker,s Angel J. Di Bilio,b I-Jy Chang,b Danilo R Casimiro,b Jay R Winkler,b Joan Selverstone Valentine,a and Harry B. Grayb aDeprtment of Chemist Y and Biochemistry, University of California, Los Angeles, California 90024 and Beckman Institute, Calijomia Institute of Technology, Pasadena, California 91125, U.S.A. Cysteines have been engineered into either the copper site or the zinc site of yeast Cu-Zn superoxide dismutase (CuZnSOD) by site-directed mutagenesis in an effort to redesign this type 2 copper protein into a type 1 copper protein. Previous spectroscopic studies [1,2] have shown that introduction of a cysteine into the distorted square planar copper site of CuZnSOD (His46Cys or Hisl2OCys) gives a copper-thiolate center with type 2 copper character, whereas introduction of a cysteine into the distorted tetrahedral zinc site (His8OCys) yields a type 1 copper. Histidines have been engineered onto the surface of the above His-to-Cys mutant proteins as well as that of the native protein (Alal21His and/or Gln123His, Lys68His and/or Thr70His). The resulting mutant proteins have been expressed in E. coli and purified to homogeneity. Modification [3] of the surface-histidine-mutant proteins by Ru(bpy)$O3 and electron-transfer studies by transient absorption spectroscopy have been carried out. The effect of cysteine on the electron-transfer properties of CuZnSOD will be discussed. 1. Y. Lu, E. B. Grab, I. A. Roe, and I. S. Valentine, J. Inorg. Biochem., 43, 184 (1991). 2. Y. Lu, E. B. Grab, I. A. Roe, and I. S. Valentine, J. Am. Chem. Sot., 114,356O (1992). 3. I.-J. Chang, H. B. Gray, and I. R Winkler, J. Am. Chem. Sot., 113, 7056 (1991).
Transcript of Electron-transfer properties of ruthenated yeast Cu-Zn superoxide dismutase mutants.
STRUCTURE/FUNCTION 109
B054 ELECTRON-TRANSFER PROPERTIES OF RUTHENATED YEAST CU-ZN SUPEROXIDE DISMUTASE MUTANTS.
Yi Lu9h b Jennifer Rennecker,s Angel J. Di Bilio,b I-Jy Chang,b Danilo R Casimiro,b Jay R Winkler,b Joan Selverstone Valentine,a and Harry B. Grayb
aDeprtment of Chemist Y
and Biochemistry, University of California, Los Angeles, California 90024 and Beckman Institute, Calijomia Institute of Technology, Pasadena, California 91125, U.S.A.
Cysteines have been engineered into either the copper site or the zinc site of yeast Cu-Zn superoxide dismutase (CuZnSOD) by site-directed mutagenesis in an effort to redesign this type 2 copper protein into a type 1 copper protein. Previous spectroscopic studies [1,2] have shown that introduction of a cysteine into the distorted square planar copper site of CuZnSOD (His46Cys or Hisl2OCys) gives a copper-thiolate center with type 2 copper character, whereas introduction of a cysteine into the distorted tetrahedral zinc site (His8OCys) yields a type 1 copper.
Histidines have been engineered onto the surface of the above His-to-Cys mutant proteins as well as that of the native protein (Alal21His and/or Gln123His, Lys68His and/or Thr70His). The resulting mutant proteins have been expressed in E. coli and purified to homogeneity. Modification [3] of the surface-histidine-mutant proteins by Ru(bpy)$O3 and electron-transfer studies by transient absorption spectroscopy have been carried out. The effect of cysteine on the electron-transfer properties of CuZnSOD will be discussed.
1. Y. Lu, E. B. Grab, I. A. Roe, and I. S. Valentine, J. Inorg. Biochem., 43, 184 (1991). 2. Y. Lu, E. B. Grab, I. A. Roe, and I. S. Valentine, J. Am. Chem. Sot., 114,356O (1992). 3. I.-J. Chang, H. B. Gray, and I. R Winkler, J. Am. Chem. Sot., 113, 7056 (1991).
