Investigation of the Structure and Kinetics of Heme-Containing Proteins Using Magnetic Circular Dichroism and Rapid-Scan, Stopped-Flow Spectroscopy
type: CreativeWork
description: Heme proteins are highly versatile with important roles in biochemistry including oxygen transport, storage, and catalysis. Work on heme proteins has often led the way for studying biological systems, and while different heme protein families have unique gene sequences, protein folding and varied roles in biochemistry, some characteristics are similar throughout all heme proteins and a variety of spectroscopic methods have been first applied to the study of heme proteins over the years. Using the two defining features of all heme proteins, the iron metal center and the tetrapyrrole macrocycle, UV-visible (UV-Vis) electronic absorption and rapid-scan, stopped-flow spectroscopies were utilized to determine reaction mechanisms and kinetics data. Due to the chromophore properties of heme-containing proteins, magnetic circular dichroism (MCD) spectrophotometry was used to determine structural aspects of different heme-containing proteins. In part one of this dissertation, two projects highlight the kinetics of reacting thiolate-ligated heme proteins with different oxo-donors. The reaction between ferric Caldariomyces fumago chloroperoxidase (CCPO) and meta -chloroperoxybenzoic acid (mCPBA) has been examined, but unlike most CCPO reactions, Compound I (Cpd I) and Compound II (Cpd II) are formed using the same reactant. Thus, the peracid is used as an oxo donor to produce Cpd I and then as a reductant to reduce Cpd I to Cpd II, and finally, Cpd II to the ferric state. In the second project, the reaction of ferric cytochrome P450cam (Cyp101) with substituted (Cl, CH3, OCH3) perbenzoic acids was studied using rapid scanning stopped flow spectroscopy. An absorption appears en route to the formation of the high-valent moiety known as Compound I that is thought to be an ferric acylperoxo heme adduct similar to the known Compound 0 of other peroxidases. Further support for this conclusion derive from additional linear Hammett correlation plots for both the rate of formation of t…
abstract: Heme proteins are highly versatile with important roles in biochemistry including oxygen transport, storage, and catalysis. Work on heme proteins has often led the way for studying biological systems, and while different heme protein families have unique gene sequences, protein folding and varied roles in biochemistry, some characteristics are similar throughout all heme proteins and a variety of spectroscopic methods have been first applied to the study of heme proteins over the years. Using the two defining features of all heme proteins, the iron metal center and the tetrapyrrole macrocycle, UV-visible (UV-Vis) electronic absorption and rapid-scan, stopped-flow spectroscopies were utilized to determine reaction mechanisms and kinetics data. Due to the chromophore properties of heme-containing proteins, magnetic circular dichroism (MCD) spectrophotometry was used to determine structural aspects of different heme-containing proteins. In part one of this dissertation, two projects highlight the kinetics of reacting thiolate-ligated heme proteins with different oxo-donors. The reaction between ferric Caldariomyces fumago chloroperoxidase (CCPO) and meta -chloroperoxybenzoic acid (mCPBA) has been examined, but unlike most CCPO reactions, Compound I (Cpd I) and Compound II (Cpd II) are formed using the same reactant. Thus, the peracid is used as an oxo donor to produce Cpd I and then as a reductant to reduce Cpd I to Cpd II, and finally, Cpd II to the ferric state. In the second project, the reaction of ferric cytochrome P450cam (Cyp101) with substituted (Cl, CH3, OCH3) perbenzoic acids was studied using rapid scanning stopped flow spectroscopy. An absorption appears en route to the formation of the high-valent moiety known as Compound I that is thought to be an ferric acylperoxo heme adduct similar to the known Compound 0 of other peroxidases. Further support for this conclusion derive from additional linear Hammett correlation plots for both the rate of formation of t…
author: Collins, Daniel Patrick
keywords: Chemistry, Physical Sciences and Mathematics, Chloroperoxidase, Cytochrome c, Cytochrome P450, Enzymes, Heme, Monooxygenase
date published: 2012-01-01T08:00:00Z
date modified: 2026-07-24T04:38:23Z
provider: University of South Carolina
publisher: Networked Digital Library of Theses and Dissertations
type: WebPage
source organization: University of South Carolina