Submitted in partial fulfilment for the Degree of Doctor of Technology: Biotechnology, Durban University of Technology, 2010. / This study involved a series of molecular dynamics (MD) simulations applied to case studies of small and medium-size polypeptides to assess the thermodynamics of their folding characteristics. Peptide folding is a complex and vital phenomenon taking place in all living systems. Bioactive conformational structures of folded peptides need to be well characterized before using them in computer-aided drug design. The computational procedure was validated on the 10-residue long chignolin-like synthetic mini-protein (CLN025). For this peptide, replica exchange molecular dynamics (REMD) calculations were carried out in explicit and implicit solvents using the generalized Born (GB)/surface area (SA) approximation with different sets of force field parameters. Following this validation procedure, case studies of the folding conformations of peptides of different lengths including the 5-residue met-enkephalin, the 27-residue pituitary adenylate-activating polypeptide 27(PACAP27) and the 28-residue vasoactive intestinal peptide (VIP) were undertaken. The latter two peptides are multifunctional hormones that mediate diverse biological functions, such as the cell cycle, cardiac muscle relaxation, immune response, septic shock, bone metabolism, and endocrine function. Results obtained indicate that when explicit water, methanol and DMSO solvents were used, it appeared that methanol (MeOH) and dimethylsulphoxide (DMSO) afforded met-enkephalin the ability to form more intra-hydrogen bonds than water, producing type I and type III β-turn structures; thus enhancing the helical conformation of the peptide. MD trajectories of longer polypeptides (VIP and PACAP27) were also populated with type I and type III β-turns, which occurred consecutively; with α- and 310-helices occurring from the middle of each peptide towards the C-terminal. Characterization of implicit solvent results, reveal that these simulations have been able to reproduce the same type of conformers obtained by experimental NMR studies published in literature, which structurally resemble the native conformation of the bioactive peptides. These conformational structures will be applied as lead agents in computer-aided drug design. One of the major achievements of this study is the ability to optimize and validate the force field parameter sets to describe the thermodynamic properties of peptide systems in an unbiased manner, a non-trivial task for even the smallest of peptides. These findings re-affirm the notion that computational methods have matured enough to model dynamic biological phenomena such as peptide folding, a feat previously thought to be impossible.

Submitted in fulfillment of the requirements of the Degree of Doctor of Technology: Chemistry, Durban University of Technology, 2011. / Understanding the dynamics and mechanism of protein folding continues to be one of the central problems in molecular biology. Peptide folding experiments characterize the dynamics and molecular mechanisms of the early events of protein folding. However, generally the highly flexible nature of peptides makes their bioactive conformation assessment reasonably difficult as peptides fold at very fast rates experimentally, requiring probing on the nanosecond time resolution. On the other hand, determining the bioactive conformation of biological peptides is a requirement for the design of peptidomimetics in computer-aided drug design. Peptides offer a unique opportunity to bridge the gap between theoretical and experimental understanding of protein folding. Therefore, the present work focuses on the exploration of the conformational space of biologically active neuropeptides with the aim of characterizing their conformational profile. Specifically, bombesin, neuromedin B (NMB) and neuromedin C (NMC), have been chosen for the current investigations. These peptides are widely distributed in the gastrointestinal tract, spinal cord and brain, and are known to elicit various physiological effects, including inhibition of feeding, smooth muscle contraction, exocrine and endocrine secretions, thermoregulation, blood pressure and sucrose regulations and cell growth. These peptides act as a growth factor in a wide range of tumours including carcinomas of the pancreas, stomach, breast, prostate, and colon. This work is intended to get some insight into the performance of different procedures used to explore the configurational space to provide an adequate atomic description of these systems. Different methodological studies involving utilization of molecular dynamics (MD), multicanonical replica exchange molecular dynamics (REMD) and simulate annealing (SA) are undertaken to explore the folding characteristics and thermodynamics of these neuropeptides. MD and REMD calculations on bombesin peptide have revealed its dual conformational behaviour never discovered before and is described in chapter 3. These results explain the known structure-activity studies and open the door to the understanding of the affinity of this peptide to two different receptors: BB1 and BB2. In the case of NMC, REMD calculations are carried out in explicit and implicit solvents, using the Generalized Born (GB) surface area, and are then complemented with two additional MD simulations performed using Langevin and Berendsen thermostats. The results obtained clearly reveal that REMD, performed under explicit solvent conditions, is more efficient and samples preferentially folded conformations with a higher content of  and γ turns. Moreover, these results show good agreement with the experimental results supporting the role of two -turns for its biological action, as reported in the literature. Finally, the results obtained from MD, REMD and SA calculations on NMB reveal that the peptide has a tendency to adopt both turns and helices suggesting its two different receptor recognizing and binding conformations during its biological action. Hence, the present work provides comprehensive information about the conformational preferences of neuropeptides which could lead to a better understanding of their native conformations for future investigations and point the way towards developing their new antagonists.