Witold Karol Subczynski, PhD, DSc
Professor of Biophysics
Department of Biophysics
Medical College of Wisconsin
8701 Watertown Plank Road
Milwaukee, WI 53226-0509
Laxman Mainali, PhD, Postdoctoral Fellow
I received my MSc in physics (1969) and my PhD in physico-mathematical sciences (1976) from Lomonosov Moscow State University (Moscow, Soviet Union) and my DSc in natural sciences (with a specialization in biophysics) in 1984 from Jagiellonian University (Krakow, Poland). I received the title of Professor of Biological Sciences (with a specialization in biophysics) from the president of Poland in 1995. Shortly after completing my training, I joined the Biophysics Department, Institute of Molecular Biology, at Jagiellonian University as a teaching/research assistant. I joined the faculty in 1977 and achieved the rank of Professor in 1995. From 1988 to 1991, I was the Chairman of the Biophysics Department, and from 1992 until 2000, the Head of the Laboratory of Structure and Dynamics of Biological Membranes. I directed 14 graduate students for their MSc degrees and supervised four students for their PhD degrees. I made my first visit to Milwaukee in 1980 and since then have been at the National Biomedical EPR Center about 50% of the time. More than 40 of my papers have been coauthored with faculty or students from the EPR Center. In 2000, I emigrated to the United States and joined the faculty of the Medical College as an Associate Professor of Biophysics.
Over the last two decades, spin-label oximetry methods have been developed and applied to study oxygen consumption and evolution in different biological and biochemical systems, as well as oxygen transport in better-defined model systems. I have been actively involved in the development and application of these methods. Although molecular oxygen is paramagnetic, direct detection of oxygen in biological systems using EPR is not possible. However, indirect methods exist in which bimolecular collisions of oxygen with paramagnetic molecules alter the resonance characteristics of these probe-paramagnetic molecules. Previously, the term "spin-label oximetry" described the application of nitroxide radical spin labels to oximetry measurements. This term should now be broadened to include other paramagnetic substances that are sensitive to collisions with oxygen because new, stable free radicals and solid-state paramagnetic probes have been introduced, especially for in vivo oximetry measurements. In my investigations, oxygen was also used as a probe to study three-dimensional molecular organization and dynamics in membranes. Finally, because oxygen and nitric oxide (NO) are paramagnetic (oxygen has a triplet ground state, while NO has one unpaired electron making it a free radical), a similar approach can be used to study NO concentration and transport in biological and model systems. It has been shown that this method, called "spin-label NO-metry," is also quantitative, giving a local NO diffusion-concentration product.
For more than 25 years I have investigated the physical properties of lipid bilayer membranes. Mainly using the EPR spin-labeling method, my research team and I have obtained unexpected results that are significant for the better understanding of the function of biological membranes. These results can be summarized as follows:
Unsaturation of lipid alkyl chains greatly reduces the ordering and rigidifying effects of cholesterol, although unsaturation alone gives only minor fluidizing effects, as observed by order and reorientational motion, and rather significant rigidifying effects, as observed by translational motion of probe molecules.
Fluid-phase model membranes and cell plasma membranes are not barriers to oxygen and NO transport.
Polar carotenoids can regulate membrane fluidity in a way similar to cholesterol.
Formation of effective hydrophobic barriers to the permeation of small polar molecules across membranes requires alkyl chain unsaturation and/or the presence of cholesterol.
Fluid-phase micro-immiscibility takes place in cis-unsaturated phosphatidylcholine-cholesterol membranes and induces the formation of cholesterol-rich domains.
In membranes containing a high concentration of transmembrane proteins, a new lipid domain is formed (with lipids trapped within aggregates of proteins) in which lipid dynamics are diminished to the level of gel-phase.
Presently, my work is focused on the study of the formation of raft domains in model and biological membranes. The cell membrane has a two-dimensional, liquid-like structure that contains domains of various time-scales and space-scales that form and disperse continuously. Rafts are this type of membrane domain, which requires lipid interactions for its formation. The long-term objective of my research is to better understand the molecular mechanisms by which rafts form and are maintained and disintegrated in biological membranes. I apply the pulse EPR spin-labeling method, discrimination by oxygen transport (DOT), for in situ studies of rafts in both model and cell membranes. This is a dual-probe saturation recovery EPR approach in which the observable parameter is the spin-lattice relaxation time of lipid spin labels and the measured value is the bimolecular collision rate between molecular oxygen and the nitroxide moiety of spin labels. The DOT method permits discrimination of different membrane domains because the collision rate between oxygen and the nitroxide moiety of spin labels (oxygen diffusion-concentration product) can be quite different in these domains. Additionally, membrane domains can be characterized by profiles of the oxygen diffusion-concentration product in situ without the need for separation.
Dr. Akihiro Kusumi, Institure for Frontier Medical Science, Kyoto University, Kyoto, Japan
Dr. Marta Pasenkiewicz-Gierula, Biophysics Department, Institute of Molecular Biology, Jagiellonian University, Krakow, Poland
Dr. Alexander N. Tikhonov, Department of Biophysics, Faculty of Physics, Lomonosov Moscow State University, Moscow, Russia
Dr. Ronald N. McElhaney, Department of Biochemistry, University of Alberta, Edmonton, Alberta, Canada
Dr. Abbas Pezeshk, Department of Chemistry, Moorhead State University, Moorhead, MN, USA
Dr. Marija Raguz, Department of Medical Physics and Biophysics, University of Split School of Medicine, Split, Croatia