Enzyme catalysis allows for enantioselective reactions to be performed at low temperatures, mild pH and atmospheric pressure. Despite these advantages, the immobilization of enzymes is crucial for the economic viability of biocatalytic processes in industry. Immobilization ensures lower losses of expensive enzymes and at the same time can improve their stability under the conditions of industrial processes that deviate greatly from the natural environment of enzymes. The implementation of biocatalytic processes in microflow systems allows for high conversions with small amounts of solvents and by-products, due to good temperature regulation, fast mass transfer and low volume. At the same time microflow systems offer the possibility of continuous operation and have received much attention lately. Nanofiber mats represent a flexible carrier for the immobilization of enzymes in microreactors. They have a large specific surface with the possibility of functionalization and at the same time, due to their high porosity, they enable efficient mass transport. In this work, expression and isolation of several amine transaminases with and without hexahistidine (His6) tags at different ends of the protein was performed and compared regarding their immobilization efficiency on nanofiber mats with different functional groups. A large loss of enzyme activity was observed when transaminases were covalently bound to the nanofiber mat, while coordinate binding via His6 tag resulted in the highest retained enzyme activity. In addition, the cofactor co immobilization on a selected nanofiber mat was successful.