Figure 2.

Synthetic biological circuits can aid in understanding of biology, improve biomanufacturing productivity, and enable disease-targeted therapy. (a) The native circuit regulating competence in B. subtilis was compared with a synthetic circuit with similar dynamics to reveal architecture-specific variability in the duration of competence and consequent differences in the consistency of transformation efficiency over large ranges of DNA concentration [71]. (b) A synthetic protein scaffold was used to increase the biosynthesis of mevalonate from acetyl-CoA in E. coli. The scaffold consists of three protein-protein interaction domains (GBD, the GTPase binding domain from the actin polymerization switch N-WASP; SH3, the Src homology 3 domain from the adaptor protein CRK; and PDZ, the PSD95/DlgA/Zo-1 domain from the adaptor protein syntrophin) in various copy numbers connected by glycine-serine linkers. Pathway enzymes (AtoB, acetoacetyl-CoA thiolase; HMGS, hydroxymethylglutaryl-CoA synthase; HMGR, hydroxymethylglutaryl-CoA reductase) were each fused to the ligands of one interaction domain and recruited to the protein scaffold [39]. P_TET, tetracycline-inducible promoter; P_BAD, arabinose-inducible promoter. (c) A targeted therapeutic circuit was constructed by inserting an RNA aptamer near an alternatively spliced exon harboring a stop codon in a three-exon, two-intron minigene fused to herpes simplex virus thymidine kinase (HSV-TK). Binding of a disease marker protein to the aptamer results in exclusion of the alternative exon, expression of a suicide gene, and killing of diseased cells [35]. P_CMV, cytomegalovirus promoter.