Publication Date


Document Type


First Advisor

Erman, James E.

Degree Name

Ph.D. (Doctor of Philosophy)

Legacy Department

Department of Chemistry and Biochemistry


Hemoproteins; Ligands (Biochemistry)


Studies of the interaction of small molecules (ligands) with heme proteins have been useful in improving understanding of structure-function relationships in heme proteins. Binding of ligands can be used to probe the structure and chemical reactivity of heme proteins. This dissertation incorporates a study of the binding of cyanide and imidazole to some mutants of cytochrome c peroxidase (CcP) and some heme sensor proteins. Binding of cyanide to a mutant of CcP in which the distal histidine was replaced with leucine was investigated. pH dependence of the binding of cyanide to this mutant was very different from the wild-type enzyme. The mutant binds cyanide anion at high pH even better that the wild-type enzyme. The binding of HCN is inhibited in the mutant due to the lack of base catalysis by the distal histidine. Distal pocket mutants of CcP in which three distal pocket residues, Arg-48, Trp-51, and His-52, were replaced with nonpolar residues were constructed. Imidazole binding to these mutants was studied in order to understand the reasons for the inability of CcP to bind imidazole. These mutants showed appreciable binding of imidazole, which may be due to the increased nonpolar nature of the heme pocket. Binding of cyanide to these mutants was very weak and slow, due to the lack of distal histidine as well as the nonpolarity of the distal heme pocket. Studies of the binding of cyanide and imidazole to heme sensors, DOS and FixLs, also emphasize the role of polarity of the heme pocket. These heme sensors have a predominantly nonpolar heme pocket. FixLs bind imidazole with fast rates. Binding of cyanide to FixLs and DOS is slow compared to CcP, which has a polar heme pocket with a distal histidine. DOS does not bind imidazole, probably due to a sixth, endogenous ligand. Comparison of the binding of cyanide and imidazole between the heme proteins included in this study showed that a polar heme pocket favors cyanide binding while a nonpolar heme pocket favors imidazole binding. pH dependence of the binding of both the ligands showed that ionizable groups in the protein influence the binding.


Includes bibliographical references (pages [288]-293).


xxii, 313 pages




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