| Proteins are the most important bio-macromolecules in organisms. Almost all of the life's activities involve proteins. Enzymes are biological catalysts in biological cells, and most of them are proteins. In the very mild conditions in the biological cells, enzymes can highly efficiently catalyze myriads of biochemical reactions, and promote organism metabolism. Biological processes, such as digestion, absorption, respiration and reproduction are all enzymatic reactions. In this meaning, enzymes are the base for a cell to live. Due to their biological significance, the mechanisms of enzymatic reactions gradually become research topics that attract lots of interests.Molecular simulation is a method that studies the relationship between the proteins structures and functions in atomic level. It provides an important tool to give a depiction of the conformation space in great detail and has become an important supplement to the experimental results. However, the quality of molecular dynamics simulation depends on the accuracy of the molecular fields it uses. QM/MM method is a very effective approach to study enzyme reaction mechanisms, by integrating quantum mechanical and molecular mechanical simulations in a whole framework. Usually, enzyme active sites are only a small portion of the system, and thus could be dealt with quantum mechanical methods. The rest of the enzyme and solution are treated with molecular mechanical methods. QM/MM methods have contributed a lot in finding out the structures of transition states and clarifying the mechanisms of enzymatic reactions.The main content of this paper is the use of molecular mechanics and quantum mechanics combined approach to study the mechanisms of the catalytic reactions of E.coli peptide deformylase (PDF) and Bacillus Thermoproteolyticus thermolysin (TLN). We use the improved NEB (nudged elastic bond) method to find out the minimum energy paths (MEP) of these reactions, and discuss why the two kinds of enzymes have different metal preferences in similar reactions. We introduce the basic concepts and methods in molecular simulations in chapter 1, and the method to determine the possible reaction paths of the enzymatic systems in chapter 2. We discuss how to calculate the activation energy of the reaction, and how to search the transition state space and intermediate space as well, for a system with thousands of degrees of freedom. By determining the transition state and the structure of enzyme active sites, we can explain the effectiveness, the specificity and the stability of the transition state structure when catalyzing a reaction. Then finally, we are able to modify these active sites or develop inhibitors to them by rational design. In order to find the minimum energy path (MEP) of the multi-step-reactions, we have developed a method based on the fast path optimization of NEB. The main difficulty in the study of the enzyme reaction mechanism is the large number of degree of unrelated freedom of the enzyme reaction system. For this purpose, we introduce some improvements to the original NEB, so that it can be applied to complex enzyme reaction systems.In chapter 3, we combine the AM1, AMBER, and QM/MM model to calculate the hydrolysis reaction paths of the PDF and TLN, on the basis of these improvements. These reactions are composed of four steps, which involve the transformation from a four-coordinated state to a five-coordinated state of the metal center with the formation and break of three chemical bonds. In particular, the PDF and TLN have almost the same reaction mechanism but completely different metal ion selectivity for catalysis. We study these two systems and compare the results, using the same model. We make no assumptions to the order of reaction steps when calculating the enzyme reaction paths. Our results agree well with other theoretical calculations results.
|
PDF Full Text Request