Honors Program Thesis (UNI Access Only)
Melisa M. Cherney
Cytochrome c--Stability; Hemoproteins--Stability; Ligand binding (Biochemistry);
Heme-thiolate proteins have been found in a variety of organisms spanning a range of functions. Most commonly, heme-thiolate proteins are used for biological catalysis or molecule sensing due to the molecular coordination scheme and supply of electrons on the central iron atom.1 The structure of heme-thiolate proteins is described later on. For this study, the property of interest for a class of heme-thiolate proteins is the protein’s ability to switch ligands and change structure. The specificity and mechanism by which the ligand switch occurs has not been well-studied. The functions of the heme-thiolate proteins we are attempting to model appear to be regulatory in nature (Table 1). 1 Understanding the mechanism of ligand switching will allow us to better understand how these proteins are capable of responding to biological conditions to regulate cellular processes. The Cherney research group is answering a basic science question rather than researching focused scientific applications, and the purpose of this research is to better understand protein regulation. The consequences of the research are important but the applications of the research are broad and currently unknown.
The ultimate goal of the Cherney research group is to build models of heme-thiolate proteins to better understand the mechanism of ligand switching. Yeast iso-1-cytochrome c (cytc) was chosen to build the models because cytochrome c has been well-studied and is easy to obtain and manipulate. So far, mutations have been made to cytc to change the stability and possible ligation schemes. The Cherney group is interested in measuring the stability of the variants using guanidine hydrochloride denaturation monitored by optical spectroscopy in order to create titration curves. By analyzing the titration curves, relative protein stabilities can be determined. 2 This allows us to compare our model proteins with other known heme-thiolate proteins and investigate what role protein stability plays in the ligand switch process. The purpose of this thesis is to probe the stability of iso-1-cytochrome c variants in order to look at the effect of various point mutations that have been introduced to create the model heme-thiolate proteins. We want to ascertain how these mutations impact the overall protein stability. I contributed to the group goal by developing a denaturation protocol and measuring the stability of two protein variants that were synthesized.
Date of Award
Department of Chemistry and Biochemistry
University Honors Designation
A thesis submitted in partial fulfillment of the requirements for the designation University Honors
1 PDF file (40 pages)
© 2015 Sarah Elizabeth Eikenberry
Eikenberry, Sarah Elizabeth, "Measuring the stability of iso-1-cytochrome c variants using heme spectra" (2015). Honors Program Theses. 176.