TT_III_L: Theoretical and Translational Medicine III. Lectures
Dr. Sziklai Dominik, Professor Dr. Kellermayer Miklós, Dr. Mártonfalvi Zsolt
Semmelweis University, Institute of Biophysics and Radiation Biology
Titin is the largest known protein of the human body. It is a component of different muscle types. It passes through the half sarcomere, connecting the sarcomeric M and Z-lines from both sides, and has many repeating and unique protein domains. Titin molecules have an important role in passive force generation during muscle elongation. This way, Titin is implementing a biological spring and a mechanical buffer. Besides the passive functions, it has an active role through its kinase domain to regulate complex metabolic mechanisms within the sarcomere. An important question about such a large molecule is its architecture within the sarcomere, and its interaction with the sarcomeric millieu. One of the mysterious sarcomeric structures, the M-line complex is a rather stable and chemically resistant entity, which exact composition is yet to be explored.
Our research concentrated on revealing the inner structure of the M-line complex and examine its topology and physical properties.
We isolated the skeletal M-complex protein from rabbit M.long. dorsi. Protein samples were put onto atomically flat and freshly cleaved mica surface. Topographical and physical measurments were carried out with Atomic Force Microscopy (AFM). We conducted nanomanipulation with the AFM tip, and tried to unwrap and cut the stable structure.
We inspected smaller and larger M-line complexes, the smaller ones have a high profile globular middle mass and less Titin is attached to them, unlike the larger ones, which are also high profiled, but the middle structure is elongated, and more Titins are attached to them. It sometimes behaves like a bulk material, but can be cut and disassembled with smaller cuts. We pulled out long elastic filaments from the complex. Topological analysis revealed a volume expansion after the nanomanipulation.
According to our results, a large and densely packed elastic molecular reservoir reside within the M-line complex. This structure is stable enough to resist the protein purification process and high ionic concentration, even to withstand detergents. It is possible to disassemble it physically with a nanotip. However controlling the parameters is not easy and the technique must be further elaborated.
Semmelweis University, Doctoral School of Theoretical and Translational Medicine