Semester

Summer

Date of Graduation

2009

Document Type

Dissertation

Degree Type

PhD

College

Eberly College of Arts and Sciences

Department

Chemistry

Committee Chair

Fred L. King.

Abstract

Protein-cisplatin interactions lie at the heart of both the effectiveness of cisplatin as an anticancer drug and side effects associated with cisplatin treatment. A greater understanding of the protein-cisplatin interactions at the molecular level can not only improve our understanding of the action of cisplatin as an anticancer drug but also inform the design of cisplatin-like agents for future use. Therefore, the interactions of cisplatin with three different model proteins were studied, which may provide theoretical basis for predicating mechanistically relevant protein-cisplatin interactions in biological fluids.;Cytochrome c (cyt c) was used as a model protein to develop a mass spectrometric approach to determine the primary binding site of cisplatin on proteins by coupling Fourier transform mass spectrometry (FT-MS) and tandem mass spectrometry (MS/MS and MS3). FT-MS permits identification of unique fragments in the adduct digest, characterized by MS/MS and MS3 to indicate that Met65 is the primary binding site for cisplatin on cyt c.;The interactions of cisplatin and transplatin with myoglobin (Mb) were compared in order to gain insights into similarities and differences between cisplatin and transplatin in their interactions with globular proteins. Prior to this research, the conditions for Mb denaturation were optimized to obtain the Mb digests for MS/MS and MS3.;Cisplatin and transplatin exhibit similar interactions with Mb. Monoadducts and diadducts were the primary adducts observed in both the interactions. MS/MS and MS3 analyses of the observed unique fragments in the digests of both the Mb-cisplatin and Mb-transplatin adducts indicate a common binding site for cisplatin and transplatin on the His116-His119 residues of Mb. This result coupled with a study of the interactions of cisplatin and transplatin with a dipeptide His∼Ser and the three dimensional (3-D) structure of native Mb shows that cisplatin and transplatin coordinate to the His116 and His119 residues on Mb.;The binding sites of cisplatin on native ubiquitin (Ub) and denatured Ub were compared in order to investigate the effect of protein conformation on the cisplatin binding sites on a protein. Results suggest that cisplatin has more binding sites on the native Ub than on the denatured Ub due to conformation effect. Three cisplatin binding sites are determined on the native Ub, in which two threonines are the primary binding site of cisplatin. On the denatured Ub, the Met1 residue is the specific binding site of cisplatin.

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