| Abstract [eng] |
Nowadays, a modern chemical synthesis faces more strict environmental regulations and, as a result, shifts toward so-called green chemistry, where biocatalytic methods play a key role. Unexplored degradation pathways of pyridine derivatives likely hide unique enzymes with the potential for biotechnological applications. This thesis discusses the investigation of new pyridine-ring-attacking oxygenases and their application in the biocatalysis of N-heteroaromatic compounds. 2-hydroxypyridine-degrading Burkholderia sp. MAK1 strain emerged as a unique biocatalyst for oxyfunctionalization of pyridine derivatives. The culprit behind biocatalytic capabilities of Burkholderia sp. MAK1 was identified as HpdABCDE monooxygenase – a member of soluble diiron monooxygenases (SDIMO) family. Since enzymes of this family have never been related to the biodegradation of N-heteroaromatic compounds, HpdABCDE monooxygenase stands out as an illustrious enzyme. Also these findings allowed for proposing a new 2-hydroxypyridine degradation pathway in bacteria. The discovery of HpdABCDE hinted that other SDIMOs may catalyse oxidation of N-heteroaromatics as well. As a result, PmlABCDEF monooxygenase was isolated. Escherichia coli whole cells bearing PmlABCDEF was introduced as a mild, regio- as well as chemoselective, efficient, and scalable method for the preparation of various aromatic N-oxides. Tailoring PmlABCDEF monooxygenase resulted in new enzyme variants exhibiting reshaped regioselectivity as well as the ability to produce dioxides and specific mono-N-oxides. |
