Applications of molecular dynamics simulation in biomolecular folding and recognition

• Main Author: Ng, Justin Tze Yang
• Other Authors: Mu Yuguang
• Format: Thesis-Doctor of Philosophy
• Language: English
• Published: Nanyang Technological University 2020
• Subjects: Science::Biological sciences::Biophysics; Science::Biological sciences::Molecular biology
• Online Access: https://hdl.handle.net/10356/137620
• Institution: Nanyang Technological University

Proteins are an important class of biomolecules essential for life. They are involved in many biological functions, such as cell signalling, transport, defence mechanisms etc. Proteins must undergo two processes, namely folding and biomolecular recognition, in order to effectively perform their functions. Molecular Dynamics Simulations have been demonstrated as a successful tool used complementary to experimental methods in the understanding of biological processes. The projects in this thesis involve the application of Molecular Dynamics simulations in the study of various aspects of protein folding and biomolecular recognition. The first project involves a study of biomolecular folding. The Human Alpha-lactalbumin Made Lethal to Tumor cells (HAMLET) complex was discovered as a tumoricidal complex of Human Alpha-lactalbumin and oleic acid cofactor in the ratio of 1:4-7. Experimental studies showed that the smallest subunit of Human Alpha-lactalbumin capable of forming tumoricidal complexes with oleic acid is its Alpha helical domains. In this project, we used Hamiltonian Replica Exchange Molecular Dynamics (H-REMD) simulations to sample the conformation space of the peptide derived from the first Alpha helical domain (Alpha1) of Human Alpha-lactalbumin in the presence of 4 oleic acid molecules. We also set up a parallel simulation system of the Alpha1 peptide alone in its apo form for comparison. In this study, we have unravelled conformational differences of the Alpha1 peptide in the presence and absence of the oleic acid cofactor. In the second project, we studied the biomolecular recognition of the protein Lipocalin Type-Prostaglandin D synthase (L-PGDS) towards the amyloid beta peptide (Aβ40). L-PGDS has been identified to contain Amyloid beta chaperone activity, with the ability to bind to Aβ40 monomers and prevent their deposition and aggregation. L-PGDS was also found to contain dis-aggregase activity towards pre-formed amyloid fibrils. Experimental evidence for the binding of L-PGDS to Aβ40 was detected in the form of residues that had significant chemical shift perturbations through Nuclear Magnetic Resonance (NMR) Spectroscopy. We aimed to construct a model detailing the interactions between L-PGDS and Aβ40. We adopted a biophysics-based approach by using H-REMD enhanced sampling and extracted high probability binding modes of the L-PGDS-Aβ40 complex through a clustering method based on high contact-probability residue pairs. Subsequent classical molecular dynamics simulations were performed to evaluate the stability of each binding mode and characterize various biomolecular interactions. This study provides the first model of L-PGDS in complex with Aβ40. We have identified and proposed several biomolecular interactions between L-PGDS and Aβ40, that can be further characterized experimentally. Such models of the interactions between L-PGDS and Aβ40 provides mechanistic insights into the biomolecular recognition functions of L-PGDS towards its binding partner. In the third project, we studied the plasticity of the Arabidopsis thaliana Profilin 3 (AtPRF3) N-Terminus Extension (NTE) and its role in enhancing binding towards Formin poly-proline (PolyP). Profilins are proteins involved in actin cytoskeleton assembly. In addition to actin binding, the association of Profilins with Formin proteins plays regulatory roles in actin cytoskeleton polymerization. Of the five Arabidopsis thaliana (At) Profilin isoforms, AtPRF3 is found to be unique and expressed with an additional intrinsically disordered NTE that contribute to enhanced binding towards PolyP. Crystal structures of AtPRF3 in complex with PolyP was unable to be resolved experimentally. Moreover, the crystal structure of AtPRF3 in its apo form with residues K31 to N37 of the NTE resolved revealed a clash of the NTE with the PolyP binding site when superimposed onto the experimentally resolved crystal structure of AtPRF2 in complex with PolyP. We used H-REMD to sample the conformation space of the AtPRF3 NTE and demonstrated the possibility of the NTE to adopt “open” and “closed” conformations to allow or occlude the binding of PolyP. Subsequently, we used classical molecular dynamics simulations and binding free energy calculations to understand the role of the NTE towards enhanced PolyP binding. We showed that the NTE can adopt multiple, non-specific, adaptive binding modes towards PolyP, consistent with experimental binding affinity measurements and the conformationally flexible nature of intrinsically disordered regions.