Experimental evidence for the role of buried polar groups in determining the reduction potential of metalloproteins: the S79P variant of Chromatium vinosum HiPIP |
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Authors: | Elena Babini Marco Borsari Francesco Capozzi Lindsay D Eltis C Luchinat |
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Institution: | (1) Food Science Laboratory, University of Bologna Via Ravennate 1020, I-47023 Cesena (FO), Italy, IT;(2) Department of Chemistry, University of Modena Via Campi 183, I-41100 Modena, Italy, IT;(3) Department of Chemistry, University of Calabria, P. Bucci I-87030 Arcavacata di Rende (CS), Italy, IT;(4) Department of Biochemistry, Université Laval, Quebec City Quebec G1K 7P4, Canada, CA;(5) Department of Soil Science and Plant Nutrition, University of Florence, P.le delle Cascine 28, I-50144 Florence, Italy, and Centro Risonanze Magnetiche, Via L. Sacconi 6, I-50019 S.to Fiorentino (Florence), Italy e-mail: luchinat@cerm.unifi.it Tel.: +39-55-2388200/4209262 Fax: +39-55-333273/4209253, IT |
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Abstract: | The amide group between residues 78 and 79 of Chromatium vinosum high-potential iron-sulfur protein (HiPIP) is in close proximity to the Fe4S4 cluster of this protein and interacts via a hydrogen bond with Sγ of Cys77, one of the cluster ligands. The reduction potential
of the S79P variant was 104±3 mV lower than that of the recombinant wild-type (rcWT) HiPIP (5 mM phosphate, 100 mM NaCl, pH 7,
293 K), principally due to a decrease in the enthalpic term which favors the reduction of the rcWT protein. Analysis of the
variant protein by NMR spectroscopy indicated that the substitution has little effect on the structure of the HiPIP or on
the electron distribution in the oxidized cluster. Potential energy calculations indicate that the difference in reduction
potential between rcWT and S79P variant HiPIPs is due to the different electrostatic properties of amide 79 in these two proteins.
These results suggest that the influence of amide group 79 on the reduction potential of C. vinosum HiPIP is a manifestation of a general electrostatic effect rather than a specific interaction. More generally, these results
provide experimental evidence for the importance of buried polar groups in determining the reduction potentials of metalloproteins.
Received: 26 April 1999 / Accepted: 24 August 1999 |
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Keywords: | Chromatium vinosum Electrostatic effects High-potential iron-sulfur protein NMR spectroscopy Reduction potential |
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