The industrial exploitation of microbes for the sustainable production of "green" chemicals and fuels contributes significantly to stepping out from the oil-based era and relentlessly pursuing a biobased, sustainable economy. Microbes have the inherent ability to produce numerous valuable compounds. However, in most cases, their natural production capacity is not enough to serve increasing market demands. Therefore, their physiological upgrade through genetic engineering is an essential prerequisite for the successful large-scale transition to microbial biorefineries. In this thesis, I developed and employed a range of cutting-edge synthetic biology technologies to enhance the biocatalytic capabilities of the industrial bacterial chassis Pseudomonas putida. I generated a set of industrially applicable platform strains and exemplified various genetic engineering tools and concepts that can be readily applied to further extend the potential of this promising bacterium.