Professor Dr. Barbara Saccà, Center of Medical biotechnology, University of Duisburg-Essen

Coupling biochemical processes in a logical order

J. Huang, A. Jaekel, J. van der Boom, D. Podlesainski, M. Elnagar, A. Heuer-Jungemann, M. Kaiser, H. Meyer, B. Saccà

In the cell, biochemical processes are coupled in a precise logical sequence to ensure that the products of the first process are used as reactants of the next process. Accordingly, individual reactions take place within specialized compartments, with transfer of matter into and among compartments being regulated by various types of gating mechanisms. The goal of this work is to emulate this control strategy in a very simple fashion using DNA origami compartments to immobilize two distinct processes in a programmable logical sequence.

The system couples the segregation/unfolding function of the p97 ATPase with the proteolytic activity of a serine protease (i.e. chymotrypsin), thus mimicking the putative coupling of the p97/proteasome machinery, which is supposed to be critical for protein homeostasis in the cell. The two events are confined within distinct DNA origami compartments, A and B, and linked in a defined spatial order, such that AB performs differently from BA. We show that compartmentalization enhances the rate of the individual reactions and that their physical separation into distinct compartments enables their otherwise incompatible coexistence in solution.

Finally, we present strategies for controlling the flow of the reaction from one chamber to the other by favoring accessibility at one side of the compartment and limiting molecular diffusion through the lateral DNA walls. We envision that this construction principle may be employed to create specialized reaction units that can be combined in a modular and programmable fashion not only to mimic naturally occurring cascades but also to perform operations not existing in nature.