Fluorescent Mensuration And Computer Simulation Of Protein Folding

This thesis mainly discussed the experimental mensurations and computer simulations of protein folding.In the experiments, the denaturalizations of BSA in various solvent environments were observed with fluorescence emission spectrum. Some phenomena were found: The relative luminescent intensity decreased while the concentration of urea increased, which illuminated the Tyr residues were exposed to the aqueous environment gradually. The location of peak in the spectrum appeared blue shift at first and then appeared red shift while the concentration of urea increased, which illuminated the conformation of the protein turned compact at first and then turned loose. Starting from native pH value, the relative luminescent intensity always decreased regardless of the pH value increased or decreased, which illuminated the conformation always turned loose. The above results accord with current theories very well. However, the location of peak in the spectrum was observed to appear blue shift while the pH value decreased. It is hard to explain this phenomenon with present theories, and more deeply researches are required.In the computer simulations, the Discrete Molecular Dynamics(DMD) method was induced. By simulating c-Crk SH3, the influences of potential parameters and heat bath methods on the thermodynamics and kinetics of system were investigated. First, it was observed that different heat bath methods can impact the simulation results. Hence, the influences of the hard-core radii and square-well radii on the results of simulations were ascertained. And the reasons were analyzed using the energy-filler theory. It is shown that the better simulation results can be acquired by modifying the potential in Borreguero and Dokholyan's model. These conclusions may provide useful indications on the more realistic Hamiltonian of DMD method as well as on the protein design.The folding mechanisms of a two-state protein c-Crk SH3 and a three-state protein Human a-lactalbumin(α-LA) were compared with DMD simulations. c-Crk SH3 only has two stable states: folding state and unfolding state. And its transition is a strong first-order like transition. Except folding and unfolding states, a-LA has another stable intermediate state. In the configuration of intermediate state, the β core is conserved as well as the a core is demolished. α-LA experiences two first-order like transitions, corresponding with the demolishment of the a core and the β core respectively. The diversities of the folding thermodynamics root in the variety of high-level structures. The results may provide useful indication to the later in-depth researches about the folding mechanisms and pathways of two-state and three-state proteins. And these results also effectively validate the viewpoint in the molecular biology: structure decides function.