Final Report Summary - ARISYS (Engineering an artificial immune system with functional components assembled from prokaryotic parts and modules) The ARISYS Project has exploited design concepts, construction hierarchies and standardization notions that stem from contemporary Synthetic Biology for the assembly and validation of an artificial all-bacterial immune-like system. To this end, an entire genetic platform has been developed based on two Gram-negative bacteria: Escherichia coli and Pseudomonas putida that affords the easy generation of Vhh domains of camel antibodies (nanobodies) upon presentation of a given antigen to a bacterial culture. The platform is composed of various separate genetic devices and on/off switches which include [i] a suite of tools for surface-display of nanobodies on the surface of either bacterium (E. coli and P. putida) encoded in their respective chromosomes, [ii] Two systems for in vivo diversification of the antigen-binding part of the nanobody through either ssDNA recombineering of the Vhh sequences with degenerate mutagenic oligos or exposure of the genomic Vhh sequences to the focused action of cytosine deaminase, [iii] a naive, ready-to use camel antibody library displayed on the bacterial surface and coupled to the diversification devices mentioned and [iv] a selection system based on inmunomagnetic capture of best antigen binders which allows automation of diversification/selection cycles. In this way, the single, and easy-to-handle—albeit heavily engineered—strains resulting from the project govern all operations that are typically scattered in a multitude of separate methods and apparatuses for antibody production.
