My Research

PhD Research: My research during graduate school in Dr. Brancaleon’s Molecular Biophysics lab at UTSA involved studying the photoinduced structural changes of Human Serum Albumin (a protein) mediated by novel photoactive derivatives of the Perylene molecule. I also used computational methods to predict the most likely binding configuration between HSA and Perylene. My overall goal was to deepen our knowledge of photoinduced mechanisms for unfolding proteins and explore the relationship between a protein’s conformation and its function. My thesis successfully accomplished three goals:

To acquire the photophysical characteristics of novel Perylene derivatives.
To show that Perylene derivatives do, in fact, interact and bind with proteins.
To show that irradiation of the Perylene while bound with protein affects the protein conformation.

One of the major possibilities offered by this research is the substitution with electron accepting or electron donating groups that bind with proteins which could modulate the photophysical properties of compounds and in particular the photoinduced electron transfer (PET) mechanism. The potential to prompt PET would provide photoactive ligands capable of modifying the local environment at the dye binding sites. The ability to control PET would be beneficial as a trigger of localized conformational changes in polypeptides and could be used to modulate the structure and function of otherwise non-photosensitive proteins. Our focus on PET stems from the fact that electron transfer (whether photoinduced or not) is a fundamental event for living systems.

Master’s Research: My master’s thesis research in Dr. Urayama’s lab at Miami University of Ohio was split into two parts. The first part was to study the effects of pressure on the conformational states of NADH (dihydronicotinamide adenine dinucleotide). Pressure is one of the fundamental environmental variables that affect states of biological systems. Pressure encountered in the biosphere can range from 1100 atm at the deepest oceans to -15 atm for water transport in trees. Understanding the effects of pressure on free NADH gives us a window into understanding life that survives in the extreme conditions on our planet. At ambient pressure, free NADH concentration is sensitive to dissolved oxygen concentration, suggesting that the free NADH concentration is more susceptible to oxidation than is the protein-bound NADH concentration. We measured the effects of pressure by characterizing the excited-state fluorescence emission of folded and unfolded states of NADH under varying pressure, and developed analytical methods for acquiring the thermodynamic properties of NADH conformational states by assuming a two-state model. The second part of my project was to construct a submicron imaging setup based on optical sectioning via structured illumination in a wide-field microscope, and to show that this setup works in an epi-illumination mode. The ultimate goal was to measure intracellular free and protein-bound NADH ratios under pressure in a model cell via spectroscopic studies combined with imaging to confirm mitochondrial localization of the NADH signal. For the imaging of structures at sub-micron scales, conventional microscopes lack sufficient resolution. Structured illumination offers an attractive means of rejecting out of focus light, can be used in real time, and can achieve resolution comparable to that of confocal and multiphoton microscopy. It can be used with incoherent light sources, and requires less data acquisition time than confocal microscopy since it is a wide-field technique. I was able to successfully section images and improve image resolution on a conventional slide mounted sample and in a capillary. These techniques were used for under pressure studies after my graduation.

Undergraduate Research: As an undergraduate student at UTPA, my work with my advisor, Dr. Dimakis, consisted of computational modeling of metalloprotein active sites. We used density functional theory, which is a quantum mechanical computational model, to study the electron structure, or more specifically the ground state electron density. It is estimated that approximately half of all proteins contain a metal atom. Thus, metalloproteins have many different functions in cells, such as storage and transport of proteins, enzymes and signal transduction proteins. Understanding the structure of metalloprotein active sites becomes even more important because of their function in infectious diseases. This research led to my first publication as an undergraduate student.