The combinations of amino acids into peptides and proteins, through peptide bonds, are the building blocks of life on earth. Their natural therapeutic properties has seen a significant increase in the application of these materials in the treatment of chronic diseases. The purest and most stable crystalline form provides structural information at the atomic level and is desirable for formulation into efficacious pharmaceutical products. Peptide crystallisation, as a good alternative to chromatographic purification, can also solve the shortcomings of traditional purification method, such as high cost, proteolytic degradation and physiochemical instability. However, peptide crystallisation still remains a major challenge due to highly flexible conformations especially in the case where water plays an integral role in the crystal structure. Glycine is the simplest amino acid and is known to play an important role in new biomimetic functional materials and biopharmaceutical research. Its hydrogen side chain makes the molecule an ideal candidate to study the effeect of chain length on the peptide solubility and crystallisation, without the effect of side chain.
The glycine homopeptides crystallisation research in this thesis includes three parts: thermodynamic properties, kinetic properties, and the relationship between peptides conformation and crystallisation. Firstly, the solubility of glycine homopeptides (glycine, diglycine, triglycine, tetraglycine, pentaglycine, and hexaglycine), amino acids with different side chains (aspartic acid, phenylalanine, histidine, and tyrosine) and their dipeptides (asp-phe, gly-asp, gly-phe, phe-phe, gly-gly, tyr-phe, gly-tyr, gly-his) in water from 278.15K to 313.15K were measured using the UV-Vis spectroscopy method and dynamic method. The modified Apelblat equation is used to correlate the relationship between solubility in water and temperature. Molecular dynamic (MD) simulation was further employed to investigate the solute-solvent interactions behind the dissolution behaviors. Moreover, the group-group interaction matrix of the SAFT-γ Mie approach was extended for the prediction of the solubility of amino acids and peptides, exploring the application of SAFT-γ Mie to biomolecular thermodynamic properties. Secondly, the classical nucleation theory was applied to the short-chain glycine homopeptide crystallisation to explore the nucleation theory of macromolecules. The nucleation parameters (nucleation rate, growth rate, interfacial surface energy, and activation Gibbs energy) were calculated based on the classical nucleation theory to explore the chain length effect on the classical nucleation mechanism of peptides, providing kinetic data to the crystallisation conditions designed for industry and modeling tools, such as gPROMS. The evidence of the non-classical nucleation phenomenon was also observed and discussed. Finally, the interaction between water and peptide molecules which can stabilize the unfolded structure of peptides and proteins was revealed, the effect of temperature and salts on the transition between unfolded and folded structure was explored, giving an inspiration to the relationship between conformation and peptide crystallisation.
The research presented in this thesis investigates the thermodynamic and kinetic properties of glycine homopeptides, as well as the flexible conformation of peptides during crystallisation, thereby providing a comprehensive strategy for designing and optimising the crystallisation process. Additionally, the research establishes a fundamental understanding of peptide crystallisation, which is extremely beneficial for future macromolecular crystallisation research.