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Co-diffusion of particles in biological systems studied using PIE-FCCS

Mallory, Donald Paul
http://rave.ohiolink.edu/etdc/view?acc_num=akron1627666937482877

Abstract Details

2021, Doctor of Philosophy, University of Akron, Chemistry.
Biophysical chemistry is an amalgamation of the three classical disciplines: physics, chemistry, and biology. Much of the biophysical chemistry field is dedicated to investigating the molecular cooperation between biochemical species like lipids and proteins that are necessary for fulfilling their cellular functions. While many such interactions have been identified and characterized, many proposed relationships have yet to be fully elucidated due to the experimental challenges involved. One important example of such interactions includes the oligomerization of G-protein coupled receptors (GPCRs); a super class of integral membrane receptors responsible for transducing extracellular stimuli into intracellular responses. Given the current prevalence of GPCR targeted therapeutics, characterizing the functional role of GPCR homo/heterodimerization is crucial for developing new pharmaceuticals. Due to the noncovalent nature of some GPCR complexes, specialized techniques capable of noninvasively measuring their prevalence is necessary for comprehensive characterization. Pulsed-interleaved excitation fluorescence cross-correlation spectroscopy (PIE-FCCS) is one of such techniques. In this dissertation we demonstrated the abilities of a time resolved fluorescence technique that measures photon emission from one or two spectrally distinct species as they diffuse through an excitation volume of known dimensions. Our first work demonstrated the use of PIE-FCCS to measure the time-dependent formation of protein corona around two separate polymeric micelle constructs as they undergo Brownian diffusion in solution. Our results indicated that of the two similarly sized PEG-based micelles, the one constructed of copolymer triblock units formed a BSA-based protein corona more rapidly. The second two projects employed PIE-FCCS on systems of lower dimensionality: cell plasma membranes. In our second work, we studied the effect of pathologic point mutations on the complexation of class A GPCR rhodopsin, which had previously been determined to participate in a monomer-dimer equilibrium when expressed in Cos-7 cells. The final work studies the influence of lipids on class A and C GPCRS. Using class C GPCRS as monomer/dimer controls, we found that modulation of either the polyunsaturated fatty acid or cholesterol membrane composition changed the dimeric properties of class A GPCRs. This work illustrates the exceptional versatility of PIE-FCCS for studying complexation of molecular species diffusing in different dimensions and chemical compositions.
Adam Smith (Advisor)
David Perry (Committee Member)
Aliaksei Boika (Committee Member)
Chunming Liu (Committee Member)
Jordan Renna (Committee Member)
179 p.
Biophysics; Chemistry

Recommended Citations

Citations

  • Mallory, D. P. (2021). Co-diffusion of particles in biological systems studied using PIE-FCCS [Doctoral dissertation, University of Akron]. OhioLINK Electronic Theses and Dissertations Center. http://rave.ohiolink.edu/etdc/view?acc_num=akron1627666937482877

    APA Style (7th edition)

  • Mallory, Donald. Co-diffusion of particles in biological systems studied using PIE-FCCS. 2021. University of Akron, Doctoral dissertation. OhioLINK Electronic Theses and Dissertations Center, http://rave.ohiolink.edu/etdc/view?acc_num=akron1627666937482877.

    MLA Style (8th edition)

  • Mallory, Donald. "Co-diffusion of particles in biological systems studied using PIE-FCCS." Doctoral dissertation, University of Akron, 2021. http://rave.ohiolink.edu/etdc/view?acc_num=akron1627666937482877

    Chicago Manual of Style (17th edition)

akron1627666937482877
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