Sulfolobus acidocaldarius – establishing a novel host for cutting-edge biotechnology at its extremes

Abstract

We live in exciting times! The importance of the once overly dominant production host Escherichia coli is dwindling, while cell culture and more exotic microbial hosts are taking over the playing field. Within this Thesis I developed fundamental methods analytic procedures and process technology alike with the capability to catalyze the adoption of the extreme thermo-acidophilic Archaeon Sulfolobus acidocaldarius in future commercially relevant bioprocesses. S. acidocaldarius grows optimally at 75 C and pH 3. It is amenable to genetic manipulation and characterized by several very unique metabolic pathways, making it a promising candidate for bioconversion processes (e.g. for upcycling of biogenic waste streams or for treatment of process wastewaters), and for the production of unique biological material (e.g. tetraether lipids used in medical applications for the formation of drug delivery vehicles). A further promising field of application is bioleaching for the economic mining of low grade ores in large scale. In general, due to its harsh growth conditions the organism is predestined for processes where sterility is difficult to achieve and/or to maintain. Difficult to sterilize batches and continuous processes which run over extended periods of time are important examples. Important prerequisites for the use of S. acidocaldarius in industrial applications, like characterization of the host s biology, development of genetic tools and clinical safety, can be considered to be met. Nevertheless, some aspects still need to be investigated in much more detail and a selection of remaining challenges were tackled within this Thesis. Firstly, for the investigation of the potential of S. acidocaldarius in the field of waste-to-value applications I performed extensive growth experiments on wheat straw and beech wood hydrolysates with wild type strains and genetically modified S. acidocaldarius mutants to utilize non-food substrate streams for the production of value-added products. Secondly, to push into pharmaceutical applications and the market of high value products I developed a defined cultivation medium adhering to Quality by Design guidelines. The medium was optimized in regards of salt and trace element composition. Complex protein hydrolysates, which can be inhomogeneous in their composition and are prone to formation of inhibiting substances under growth conditions, were replaced by a combination of sodium glutamate and citric acid. Thirdly, for the assessment of cell viability, a critical parameter during process development and control, fluorescence based assays were developed. Finally, I determined critical process parameters during the cultivation of S. acidocaldarius for the direct manipulation of the organisms membrane composition. Thereby it is possible to generate a tailor-made pattern of industrially relevant lipids that might be recoverable to an increased yield via a greatly facilitated downstream procedure.