Fig. 1.

Libraries for promoter and protein evolution inP. putida. (A) Alignment of the evolved PcaU promoter (pPcaU1.1) with an E. coli adapted promoter (pPcaU1). Positions randomized to create the library are shown below the alignment. The PcaU operator is only partially shown. (B) Alignment of homologous PobR and PcaU sequences in the region with high sensitivity to mutagenesis in PobR. The D139N mutation in a PobR variant exhibited an enhanced contrast ratio (Jha et al., 2016), and randomization was carried out on PcaU at analogous positions. The corresponding region in PcaU1.2 is aligned to show TD→GY mutation. (C) Homology model of PcaU inducer binding domain as a dimer and docked with PCA (shown in spheres), showing spatial orientation of T147 and D148 (shown in sticks).