A New Strategy For The Improvement Of Biological Thermostability By Biomineralization

Biomineralization refers to the processes by which living organisms deposit minerals, and it links the soft organic material and the hard inorganic materials on the earth. Along with the evolution, almost all organisms have evolved complicate and delicate biomineralization mechanisms to control the biomineral deposition, For example the magnetotactic bacteria deposit iron oxide in enclosed organic stealths, diatoms and sponges control the formation of the macroscopic silica structures, mollusks form calcite crystals in their shells, vertebrates generate apaptite in bones and enamels. Generally, organic macromolecules such as protein and glycoprotein play an important role in regulating these biomineralization processes. The polymers with similar residues and biomimetic polypeptides are the favorable substitution in the biomimetic studies, as they are easier to be obtained and controlled. Recently, they have been broadly adopted in the material synthesis and organism biomineralization.Inspired by these biomineralization phenomena, functional inorganic materials can be introduced to living organisms in a biomimetic way to improve their performance in harsh environments. Organic biomolecules, such as protein and glycoprotein, play a key role in the control and regulation of biomineralization. Due to the complex of proteins, biomimetic polypeptides are widely used in preparing functional materials and studying organism mineralization. By introducing an inorganic exterior on organisms, we can confer them some new characteristics, such as themostability and water retention capability. Yeast, as a typical unicellular eukaryote, is a good living model in the research. Virus, which widely be used in material construction, cargo delivery and vaccine development due to its composition and structure simplicity, is also a good model. Futhermore, virus can feasibly be operated to display nucleating peptides by genetic engineering because viral genome is simple.In this dissertation, we confer the organisms such as yeast and virus vaccine an inorganic exterior by biomimetic mineralization under mild condition, which significantly improved the organisms’thermostabiltiy and water retention capability. This study mainly focuses on the development of liquid thermostable vaccines by using a biomineralization strategy. This dissertation can be divided into five chapters: Chapter1. The introduction mainly composed of an overview of biomineralization, biogenic minerals such as silica, calcium phosphate (CaP), calcium carbonate, iron minerals and amorphous phrase; the roles of protein, polymer and peptide in the control of calcium phosphate and silica biomineralization; biomimetic principle and applications; yeast cell encapsulation and mineralization, cell surface modification with polymers and inorganic materials; the chemical and biological modification of viral capsid, and functional nanomaterials by using virus as the template; the importance of vaccines and their thermostability, approaches to thermostable vaccines; the strategy and aims of this study.Chapter2. We conferred the yeast cell an inorganic calcium phosphate or silica shell by layer-by-layer method, and further studied the implications of this modification. The silica nanocoat could improve the thermostability and water retention capability upon drying condition.Chapter3. A eggshell-like calcium phopahte shell was introduced on Japanese encephalitis virus by in situ biomineralization. This eggshell-like exterior significantly improved the thermostability of virus vaccine without impairing its biological characteristics. This study provides a novel approach to thermostable vaccine development in developing countries.Chapter4. Using the human enterovirus type71(EV71) vaccine strain as a model, we suggest a combined, rational design approach to improve the thermostability and immunogenicity of live vaccines by self-biomineralization. The biomimetic peptides are rationally integrated onto the capsid of EV71by reverse genetics so that calcium phosphate mineralization can be biologically induced onto vaccine surfaces under physiological conditions, generating a mineral exterior. This engineered self-biomineralized virus was characterized in detail for its unique structural, virological and chemical properties. Analogous to many exteriors, the mineral coating confers some new properties on enclosed vaccines. Such a combination of genetic technology and biomineralization provides an economic solution for current vaccination programs, especially in developing countries that lack expensive refrigeration infrastructures.Chapter5. We showed a feasible strategy to produce thermostable liquid vaccines by introducing a protective hydrated amorphous silica exteriors to them with tunable in situ silicification. The hydrated amorphous silica exterior does not impair the original biological behaviors of virus vaccines, but confers a significantly improved thermostability on them at a wide range of temperatures. After silicification, the storage period of Poliovirus and EV71virus at room temperature can be extended by about7-fold and10-fold respectively, realizing the storage of these vaccines at room temperature for more than one month. Silica exteriors confined nearby water molecules through hydrogen bonds formed between water and hydroxyl groups, and therefore decreased the exchange of ionic and hydrogen bonding between solution and interior vaccine. The technology may pose great impact on current vaccination program by providing an economic strategy for producing liquid thermostable vaccines. Based on these results, we suggest a new concept that the silica achors can improve organisms’thermostability. The study also provides a reasonable explanation why the nature organisms often choose amorphous silica as the biominerals.Chapter6. We briefly summaried the studies in this dissertation. Our studies clearly demostrate that the biomimetic mineralization strategy can be successfully applied in improving organisms’ themostability, and hold great promise in the development of liquid thermostable vaccines. We further disscussed how to better applied this functional materials based biomimetic strategy to improve the biologicals’ applications in future.