Study On The Interactions Between Protein And Ligand With Molecular Modeling Approaches

With the completion of the human genome project, the post-genomic era, which mainly focuses on functional genomics and proteomics, has come. Proteins are important components of organisms,playing indispensable role in many kinds of life processes. The relationship between structure and function of protein is one of the pioneering and hot issues in the current life science researches. The interactions and recognition between protein and ligand is the main way through which does the protein exert its biological functions, playing crucial role in various life activities, such as gene regulation, signal transduction and immune response. Therefore, the study of the interaction mechanism between protein and ligand is of great significance for understanding the regulatory mechanisms of organisms, and also providing necessary theoretical basis for the research and development of new drugs.Since determining structure of protein complex with experimental approach is still challenging, recently, with the continuous progress in computers'processing ability and the rapid development and extensive application of theoretical simulation, molecular modeling methods, such as molecular dynamics (MD) simulation, molecular docking and free energy computation, have become important tools for exploring the interaction process of protein with its ligand. Molecular modeling not only enables us to understand and reveal the essence of life phenomena in the level of molecule, subunit or even atom, but also provides strong theoretical support to experimental results. With the improvement of molecular modeling theory and technology advances, molecular modeling methods are increasingly being used in the research of protein structure-function relationship, protein-ligand mutual recognition, as well as rational drug design.Exchange and transport of substances between environment and the inside of a cell is vital for the cell to maintain its normal life activity. Periplasmic binding protein (PBP) can recognize and bind substrate specifically, and is responsible for its delivery to the corresponding receptors or transporter proteins in the cell membrance to help regulate the translocation of nutrients or initiate chemotaxis. Study on the binding and unbinding processes of PBPs is very helpful for us to understand the substances exchange and transport process. As exemplified by the B12 binding protein BtuF and the two heme binding protein ShuT and PhuT, the structure-function relationship and the mutual recognition mechanism of the third class PBPs are thoroughly discussed in Chapter 3 and 4, respectively.Since was first discovered in June 1981, Acquired Immune Deficiency Syndrome (AIDS) speeded rapidly on a global scale and soon become a serious threat to human life and health. In the year of 1983, Barre-Sinousi and colleagues first identified the human immunodeficiency virus type 1 (HIV-1) as the pathogens of AIDS. HIV-1 integrase (IN) is an important target for the design and development of novel anti-HIV drugs. In the life cycle of HIV, IN is responsible for the integration of viral DNA into the host chromosome. Consequently, the study on the recognition mechanism of IN with inhibitors is important for the design and modification of the anti-HIV drugs. In Chapter 5 of this dissertation, the binding modes of HIV-1 IN with the coumarin-containing IN inhibitor NSC158393 is explored, and the inhibitory mechanism of this inhibitor is explained. In Chapter 6, a complex structure of HIV IN and DNA was constructed and this complex model can be used in virtual screening or other receptor-based drug design. The main content of this dissertation includes the following two parts:1. Study on the mechanism of the third class preriplasmic binding protein with molecular modeling(1) Study on the mechanism of the BtuF periplasmic binding protein for vitamin B12 with molecular modelingIn this work, the steered molecular dynamics simulation (SMD) and other molecular modeling methods were employed on the B12-bound BtuF to investigate the energetics and mechanism of BtuF, and a potential of mean force along the postulated vitamin B12 unbinding pathway was constructed through Jarzynski's equation. The large free energy differences of the postulated B12 unbinding process inticates that the B12-bound structure is in a stable closed state and some conformation changes may be necessary to the B12 unbinding. From the result of the principal component analysis, we found that the BtuF-B12 complex shows clear opening-closing and twisting motion tendencies which are consistent with the so-called"Venus-flytrap"mechanism taken by the first and the second periplasmic binding proteins (PBPs). The intrinsic flexibility of BtuF was also explored. Based on these results, it is found that the Trp44-Gln45 pair situated at the mouth of the B12 binding pocket may act as a gate in the B12 binding and unbinding process. (2) Study on the mechanism of the bacterial periplasmic heme binding proteins ShuT and PhuT with molecular modelingBy performing a series of long time MD simulations on the ShuT and the PhuT, the dynamics properties and functions of the two PBPs were investigated. The flexibilities of the two proteins are much higher than previously assumed. Through monitoring the distance changes between the N/C-terminal domains of ShuT and PhuT, it was found that the two PBPs, when in unbound state, exhibit obvious opening–closing motions. Based on the results of the domain motion analysis, large scale conformational transitions were found in all heme-free runs of ShuT and PhuT, hinting that the domain motions of the two PBPs may be intrinsic. On the basis of the results of the principle component analysis, distinct opening–closing and twisting motion tendencies were observed not only in the heme-free, but also in the heme-bound simulations of the two PBPs. The Gaussian network model was applied in order to analyze the hinge bending regions. The most important bending regions, which may act as hinges, of ShuT and PhuT are all located around the midpoints of their respective linker. In conclusion, the two heme binding protein ShuT and PhuT may also take the so-called"Venus-flytrap"mechanism during the process of capturing and releasing their substrates.2. Study on the interactions between HIV-1 integrase and inhibitors with molecular modeling(1) Study on the inhibitory mechanism and binding mode of the coumarin compound NSC158393 to HIV-1 integrase with molecular modelingIn this work, the binding modes of the wild type IN core domain and the two mutants (W132G and C130S) with the 4-hydroxycoumarin compound NSC158393 were first obtained by using the"relaxed complex"molecular docking approach combined with MD simulations, and then the Molecular Mechanics Generalized Born Surface Area (MM-GBSA) method was employed to evaluate the key residues in the binding of HIV-1 IN with NSC158393. In principle, NSC158393 binds the 128-136 peptides of IN, however, the specific binding modes for the three systems are various. According to the simulation results for the complex of the wild type IN and the inhibitor, NSC158393 can not only diminish the stability of the IN dimmer and disturb the formation of IN multimers, but also highly affect the flexibility of the functional 140-149 loop. Comparing with the wild type IN, NSC158393 can not effectively affect the flexibility of the functional 140-149 loop in the two mutants, and it can not interfere with the stability of the IN monomers either. Therefore, NSC158393 can not efficiently inhibit the integration process catalyzed by the two mutants. Three key binding residues (W131, K136, and G134) were discovered by energy decomposition calculated with the MM-GBSA method. Characterized by the largest binding affinity, W131 is likely to be indispensable for the binding of NSC158393.(2) Study on the binding modes of the HIV integrase-DNA complex to some integrase inhibitors with molecular modelingIn the current work, a model for complex of IN and viral DNA was built by combining experimental data with the results of SMD simulation. According to the results of molecular docking, the inhibitors of the second group share a similar binding model with those of the first group, though they have no common scaffold. This suggests that these inhibitors may share a similar inhibitory mechanism. In principle, these compounds interact the Mg2+ ion with their DKA-like moieties and make hydrophobic interactions with a pocket around the functional 140-149 loop with their halogenated-benzene moieties. Finally, the interactions between IN and one of these inhibitors were explored to understand their binding modes. Some key residues for ligand binding, such as T66,D116,F139 and I141, are identified according to the calculation on the interactions, and all of which have been proved to be important for DNA recognition or the catalytic activity of IN. All main findings of this work are in good accordance with previous experiment or simulation works and the newly built model of the IN-DNA complex is helpful for the subsequent research on the design of IN inhibitors.