Design and development of protein-polymer assemblies to engineer artificial organelles

Molecular engineering by means of the development of artificial organelles that can be implanted in cells to treat pathological conditions or to support the design of artificial cells has been regarded as a major goal in medical research. In order to achieve this goal, active compounds, such as enzymes, proteins and enzyme-mimics were encapsulated or entrapped in polymer compartments to create highly active hybrid nanoreactors. However, probably due to the early stage of research, only few of these nanoreactors were active in cells, and none was proven to mimic a specific natural organelle. In the present thesis, the engineering of polymeric vesicles for the development of artificial organelles mimicking natural peroxisomes is accomplished. In addition, a detailed study of the permeabilisation of polymeric vesicle membranes by incorporation of ion-channels is elaborated. The detailed study of metal-functionalised polymeric vesicle membranes for targeting approaches \'96 an important factor for site specific action of e. g. artificial organelles \'96 gives a further key aspect. These three parts of fundamental research play a key role in further development of advanced nanoreactors to be used as specific artificial organelles. In the first study, we designed systems that contained two different highly active antioxidant enzymes, which worked in tandem in polymer nanovesicles. The membrane of the nanovesicles was specifically permeabilised by the incorporation of natural channel proteins and the functional nanoreactor was optimised with regard to properties and function of natural peroxisomes. These non- toxic artificial peroxisomes combat oxidative stress in cells after cell- uptake, which prolong cell life-time and which can be regarded as the first instance of treatment of various pathologies (e.g. arthritis, Parkinson\'92s, cancer, AIDS) effectively controlled by conditions of oxidative stress. The second study proofs the successful incorporation of small ion-channels in the vesicle membrane. These ion-channels are specifically permeable for monovalent cations, for example to design a pH-sensitive artificial organelle, where the active compound can be switched on/off respectively, depending on the conditions of the environment. The third project focussed on the decoration of metal-functionalised polymersomes with a small model peptide, a His6-tag, in order to characterise the change of the metal coordination site upon binding of the model peptide. This provided important information about the binding behaviour of ligands in processes of molecular recognition, a key principle in nature. Understanding of specific and efficient binding will allow to further target the polymersomes to specific locations to either release their content, such as drugs, or to act as an artificial organelle.