Meng, Rong
(2006)
Studies of Copper (II) Complexes of Bioactive Peptides.
Doctoral Dissertation, University of Pittsburgh.
(Unpublished)
Abstract
Separation-based determinations of peptides hold the promise of measuring the concentrations of many peptides and their metabolites. However, typical chromatographic methods bear disadvantages in their selectivity and/or sensitivities. Thus, means for lowering detection limits are needed. The biuret reaction, the coordination of Cu(II) and peptides in basic solutions, provides a sensitive detection method based on the reversible Cu(III)/Cu(II) electrochemistry in peptide-bonded states.Thyrotropin-releasing hormone (TRH) is an important neuropeptide that undergoes the biuret reaction. The cationic form of TRH is not retained in a typical reversed-phase column. We have modified the common acidic mobile phase for peptide separation with a surfactant to retain the cationic TRH. A surfactant-preconditioned packed-bed capillary serves as the sample loop to preconcentrate TRH through sequential loading. The preconcentration has improved the detection limit for TRH 50-fold in the capillary HPLC-EC system. TRH lacks the N-terminal amine and C-terminal carboxylate that are usually required in the biuret reactions of Cu(II) and tripeptides. The unique structure of TRH affects the electrochemical behavior of the complex. We have utilized various electrochemical and spectroscopic techniques to determine the stoichiometry of the complex and to identify the binding sites in the peptide. Our data reveal that the interactions between Cu(II) and TRH are also possible under physiological pH. The unusual structure of TRH complicates the electrochemistry of Cu(II)-TRH complex. Multiple chemical reactions accompany the redox processes. We have investigated the kinetics of the coupling reactions using a rotating ring-disk electrode. The results suggest deprotonation of the ligand and inter-molecular interactions between complexes. Using microelectrode voltammetry, we have discovered that the octarepeat peptide of the prion protein is involved in Cu(II)/Cu(I) electrochemistry at physiological pH. This observation is relevant to the copper toxicity: Cu(II) may be reduced in vivo to Cu(I), which reacts with hydrogen peroxide to produce damaging oxidative radicals. The octarepeat peptide stabilizes Cu(I) by altering its redox potentials. Thus, the prion protein may have a function of anti-copper-toxicity.
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Details
| Item Type: |
University of Pittsburgh ETD
|
| Status: |
Unpublished |
| Creators/Authors: |
|
| ETD Committee: |
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| Date: |
2 June 2006 |
| Date Type: |
Completion |
| Defense Date: |
27 April 2006 |
| Approval Date: |
2 June 2006 |
| Submission Date: |
24 April 2006 |
| Access Restriction: |
No restriction; Release the ETD for access worldwide immediately. |
| Institution: |
University of Pittsburgh |
| Schools and Programs: |
Dietrich School of Arts and Sciences > Chemistry |
| Degree: |
PhD - Doctor of Philosophy |
| Thesis Type: |
Doctoral Dissertation |
| Refereed: |
Yes |
| Uncontrolled Keywords: |
Copper; electrochemistry; HPLC; peptide; prion; TRH |
| Other ID: |
http://etd.library.pitt.edu/ETD/available/etd-04242006-214552/, etd-04242006-214552 |
| Date Deposited: |
10 Nov 2011 19:41 |
| Last Modified: |
15 Nov 2016 13:42 |
| URI: |
http://d-scholarship.pitt.edu/id/eprint/7592 |
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