A fluorescence quenching study of the pH dependence of binding between horse heart cytochrome c and cytochrome c peroxidase

Author

Li Lin

Publication Date

1995

Document Type

Dissertation/Thesis

First Advisor

Erman, James E.

Degree Name

M.S. (Master of Science)

Legacy Department

Department of Chemistry

LCSH

Cytochrome c; Cytochromes; Hydrogen-ion concentration; Peroxidase

Abstract

The fluorescence quenching technique provides a method for studying the binding between horse heart cytochrome c and protoporphyrin cytochrome c peroxidase. The effect of pH on the binding was studied. The number of binding sites have long been in contention. Based on a 1 : 1 complex formation model, three cases for analyzing the binding using the fluorescence quenching technique were investigated in the range pH 4.0 to pH 8.0. Two models were established. Under the experimental condition, the one binding site model worked well. The second binding site was not observed by using fluorescence quenching. The equilibrium dissociation constants, which are a measure of the binding, were determined. They were less than 1 nM above pH 5.2, the isoelectric point of the enzyme, in 10 mM phosphate buffer, and increased significantly below pH 5.2 in acetate buffer. Binding between cytochrome c and porphyrin cytochrome c peroxidase was much stronger above the isoelectric point than that below the isoelectric point. The effect of apocytochrome c peroxidase on the binding with horse heart cytochrome c was also investigated. It was found that apoCcP and pCcP have the same affinity when binding with cytochrome c. Methods of calibration of extinction coefficients of porphyrin cytochrome c peroxidase for different pH, ionic strength, and wavelength were established.

Comments

Includes bibliographical references (pages [96]-100)

Recommended Citation

Lin, Li, "A fluorescence quenching study of the pH dependence of binding between horse heart cytochrome c and cytochrome c peroxidase" (1995). Graduate Research Theses & Dissertations. 304.
https://huskiecommons.lib.niu.edu/allgraduate-thesesdissertations/304

Extent

x, 117 pages

Language

eng

Publisher

Northern Illinois University

Rights Statement

In Copyright

Rights Statement 2

NIU theses are protected by copyright. They may be viewed from Huskie Commons for any purpose, but reproduction or distribution in any format is prohibited without the written permission of the authors.

Media Type

Text