(175ax) Transcriptional Regulation of Engineered Living Materials

Extracellular electron transfer (EET) is an anaerobic respiration process in bacteria that couples the oxidation of carbon to the reduction of extracellular metals. By redirecting the electron flow to a suitable redox catalyst, we have been able to control two types of redox catalysis via a bacterial interface: Atom Transfer Radical Polymerization (ATRP) and Cu(I)-catalyzed Alkyne-Azide Cycloaddition (CuAAC). By directing the electron flux to facilitate the cross-linking of long chain polymers we can control material stiffness as a function of conversion. In the EET-capable bacteria S. oneidensis, the protein pathway responsible for electron transform is the well-defined Mtr-pathway. Using transcriptional Buffer gates originally developed to control the output of fluorescent proteins, we were able to control the expression of the terminal Mtr-pathway protein MtrC via multiple orthogonal promoters. This enabled control of material stiffness as a function of mtrC expression. Then we were able to increase the complexity of transcriptional logic by placing multiple nested-promoter pairs for Boolean (2-input, 1-output) to create logical materials that selectively stiffen as a result of the presence or absence of multiple inducers. As a proof-of-concept application, this Boolean logic system was used to create hydrogels to serve as a scaffold for human fibroblast cells. Depending on the logic programed into the bacterial cells, the fibroblast cell morphology could be controlled utilizing the same logic. This engineered living materials base allowed for control over fibroblast cell morphology without requiring direct engineering of the fibroblast cells.