Kabagambe, Benjamin
(2008)
SPECTROSCOPIC INVESTIGATION OF PROTEINS.
Master's Thesis, University of Pittsburgh.
(Unpublished)
Abstract
Apomyoglobin is obtained from pure myoglobin by extracting out the heme group. UsingUV-visible spectrometry we are able to monitor complete removal of heme group. Thisextraction initiates the disruption of myoglobin's tertiary conformation. Apomyoglobin consistsof 8 α-helices labeled A through H. These helices unfold when the protein is subjected to pHdecrease. It is more compact at about pH 7 and unfolds as we change to more acidicenvironment. At about pH 2 we have A, G, H core with the rest of the helices unfolded. At pHvalues slightly lower than 2, A, G and H helices are also unfolded. When the protein is notdenatured, the refolding process is done by changing the pH towards neutral. At pH 2, G and Hhelices refold and at pH 4 A-helix refolds as well. We have been able to label A-helix withprotons and the rest of the protein with deuterium using H2O and D2O respectively. We havethen used temperature change to initiate the unfolding of protonated A-helix. When dissolved inD2O, the protonated A-helix at elevated temperatures, exchanges its N-H protons to N-Ddeuterons. Temperatures were elevated from 0 °C to 80 °C and changes in Am III peak intensitieswere observed using UV resonance Raman spectroscopy. These peak changes were quantifiedand used to calculate the number of N-H bonds present between 0 °C and 80 °C. The number ofN-H bonds decreased with increase in temperature indicating the unfolding of A-helix.
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Details
| Item Type: |
University of Pittsburgh ETD
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| Status: |
Unpublished |
| Creators/Authors: |
|
| ETD Committee: |
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| Date: |
17 January 2008 |
| Date Type: |
Completion |
| Defense Date: |
12 October 2007 |
| Approval Date: |
17 January 2008 |
| Submission Date: |
23 October 2007 |
| 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: |
MS - Master of Science |
| Thesis Type: |
Master's Thesis |
| Refereed: |
Yes |
| Uncontrolled Keywords: |
Apomyoglobin; Spectroscopy; UV Resonance Raman |
| Other ID: |
http://etd.library.pitt.edu/ETD/available/etd-10232007-094038/, etd-10232007-094038 |
| Date Deposited: |
10 Nov 2011 20:03 |
| Last Modified: |
15 Nov 2016 13:50 |
| URI: |
http://d-scholarship.pitt.edu/id/eprint/9507 |
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