Study On The Interactions Between Protein Receptor And Ligand Via Molecular Modeling Approaches

With the completion of Human Genome Project (HGP), the post-genomic era focusing on the functional genomics and proteomics researches has already come. Studies on the relationship between protein structure and function, and its actual application are important parts of the proteomics research. Protein molecules play crucial roles in the life activities through recognition and interactions with ligand molecules. These interactions are the necessary prerequisites for gene regulation, signal transduction, immunoreaction etc. Therefore, the studies of the interactions between protein receptor and ligand are important for understanding biological regulation mechanism, and cellular structure and function, which will also provide the theory foundation for designing and discovering new drug targets. The study of the interactions and recognition between protein and ligand is one of the pioneering and hot themes in life science field all the time.It is difficult to determine a protein complex structure through experimental methods. Recently, with the continuous progress of computers'processing ability, as well as the rapid development and extensive application of theoretical simulation, molecular modeling methods, such as molecular dynamics (MD), molecular docking and free energy computation, have become important tools for exploring the interaction mechanism of protein receptor with ligand. Exchange and transport of substances between inside and outside of a cell is vital for the cell to maintain normal life activity. Specific recognition and binding of periplasmic binding proteins with their ligands play a key role for the transport of the ligands. Studying the specific binding process of binding protein with ligand and exploring the structural basis for their binding are of great help for understanding this life process. The first part of this dissertation took one of the periplasmic binding proteins, Glutamine-binding protein (GlnBP), as an example to explore the binding free energy, the key residues and the hinges function during the process of GlnBP binding with its ligand Gln.The prevalence of AIDS dramatically threatens human life health, which makes the drug design against AIDS as a hot field and many countries are spending huge money on it. Human immunodeficiency virus (HIV) integrase (IN) is an important target for designing and developing the novel anti-HIV drug. In the life cycle of HIV, IN aids the integration of viral DNA into the host chromosome. Thus, the study of the effective recognition mechanism of IN with viral DNA is important for designing the anti-HIV drug against IN. The interactions of HIV IN with its different ligands (i.e. small molecular, peptide and DNA) were studied in the second part of this dissertation. Some sound interpretations were provided for a series of important biological topic, such as IN drug resistance, inhibiting mechanism of the small peptide inhibitor, and the binding mode of IN with viral DNA.The main content and creative items in this dissertation include two aspects as follows:1. MD Simulations and Free Energy calculation on GlnBPGlnBP is one of the ligand-specific periplasmic binding proteins in the Escherichia coli permease systems and plays an important role in transferring Gln from the periplasmic space to the cytoplasmic space. In this dissertation, the interactions of the key residues in GlnBp with the ligand Gln and the functional difference between the two hinges in GlnBp were evaluated through MD simulation sampling, and the binding free energy of GlnBp with Gln was calculated by the MM-PBSA method. The results show that the main impetus to bind Gln lies in the van der Waals'(VDW) interactions of Gln with Phe13, Phe50, Thr118 and Ile69, as well as the electrostatic interactions among Arg75, Thr70, Asp157, Gly68, Lys115, Ala67, His156 and ligand Gln. The hinge region 85～89, whose fluctuation is larger than the hinge region 181～185, has a more flexibility. This can provide a structural basis to bind ligand Gln, while the main function of the hinge region 181～185 was proposed to restrict ligand Gln in the pocket of GlnBp. It is found that the binding free energy between GlnBP and ligand Gln, which was predicted with the MM-PBSA method, agrees well with the experimental data.2. Study on the interactions between HIV-1 IN and its ligands via molecular modeling(1) The drug resistance and the binding mode of HIV-1 IN with L-Chicoric acid (LCA) inhibitorThe binding mode of the core domain of the wild type IN core domain and its G140S mutant with LCA inhibitor were investigated by using multiple conformation molecular docking and MD simulation. Based on the binding modes, the drug resistance mechanism was explored for the G140S mutant of IN with LCA. The results indicate that the binding site of the G140S mutant of the IN core domain with LCA is different from that of the core domain of the wild type IN, which leads to the partial loss of inhibition potency of LCA. The flexibility of the IN functional loop region and the interactions between Mg2+ ion and the three key residues (i.e. Asp64, Asp116, Glu152) stimulate the biological operation of IN. The drug resistance also lies in several interaction changes, such as the repulsion between LCA and E152 in the G140S mutant core domain, the weakening of K159 binding with LCA and Y143 pointing to the pocket of the G140S mutant.(2) The recognition and inhibition molecular mechanism of HIV-1 IN by EBR28 peptide inhibitorRecently synthesized 12-mer peptide EBR28, which can strongly bind to IN, is one of the most potential small peptide leading compounds inhibiting IN binding with viral DNA. However, the binding mode between EBR28 peptide with HIV-1 IN and the inhibition mechanism remain uncertain. The binding modes of EBR28 with HIV-1 IN monomer core domain (IN1) and dimmer core domain (IN2) were investigated by using molecular docking and MD simulation methods. The results indicate that EBR28 binds to the interfaces of the IN1 and IN2 systems mainly through the hydrophobic interactions with theβ3,α1 andα5 regions of the proteins. Based on this binding mode, the binding free energies for IN1 with a series of EBR28 mutated peptides were calculated with the MM/GBSA model, and the correlation between the calculated and experimental binding free energies is obvious (r = 0.88). Thus, the validity of the binding mode of IN1 with EBR28 was confirmed. The inhibition mechanism of EBR28 was explored by the essential dynamics (ED) analysis, energy decomposition and the mobility of EBR28 in the two docked complexes. The proposed inhibition mechanism is represented that EBR28 binds to the interface of IN1 to form the IN1_EBR28 complex and prevents the formation of IN dimmer, and finally leads to the partial loss of binding potency for IN with viral DNA.(3) The conformational changes and binding mode of HIV-1 IN with viral DNAThe specific binding mode between IN and its substrate 27 bp segments of viral DNA was obtained via the molecular docking method. The results show that the key residues for IN dimer binding with viral DNA are Lys14, Arg20, Lys156, Lys159, Lys160, Lys186, Lys188, Arg199 residues in the chain b and Lys219, Trp243, Lys244, Arg262, Arg263 residues in the chain A. The explanation for the minimum length of 15 bp viral DNA to activate IN was given based on the docked complex structure. Through the binding energy analysis, it is found that non-polar interactions are the principal factor favoring the binding of IN with DNA; whereas, the stable association of viral DNA with the key residues are mainly driven by polar interactions.Based on the binding mode obtained by molecular docking, the change of motive mode, correlative movement and viral DNA conformational change were explored with MD simulation and statistical methods. Then, solvent effect during the association IN dimer with viral DNA was analyzed briefly. The result shows that viral DNA can be divided into five regions (i.e. non-binding region, high-affinity region 1, weak-affinity region, high-affinity region 2 and reaction region) according to the binding ability. After the association of viral DNA with IN dimer, there are some significant changes in both the motive mode and the cooperative movement for each system. Compared with viral DNA before binding with IN dimer, some big conformational changes occurred for the bases in the binding region other than the non-binding region for viral DNA complexed with IN. The obvious deviation from standard B-DNA in the viral DNA main chain of the complex and the broading of the minor groove in the binding site are both the fundamentals for the recognition basis of viral DNA with IN dimer. Through analyzing the hydrogen bonds formed by water in the interface between IN dimer and viral DNA, it is found that water molecules play an important role in the recognition between IN and viral DNA.