Figure 4.

The enzymatic activity of the different recombinant proteins in the centrifuged culture medium and cell extracts. The production of EcPGA in the strain with the expression cassette P_KAR2(FL)-sfGFP (A) and in the control strain (B), CaLB (C), and TlXynA (D) was assessed as enzymatic activity (U L⁻¹) both in the culture medium, i.e., in the supernatant (dark green, dark gray, dark blue, dark red for EcPGA, EcPGA, CaLB, TlXynA, respectively), as well as in the cell extracts, i.e., inside the cells (light green, light gray, light blue, light red for EcPGA, EcPGA, CaLB, TlXynA, respectively). The concentrations were recalculated to the volume of the culture broth. The activities of the extracellular (secreted) enzyme were related to the mass of total extracellular protein (U g${}_{\text{tot}.\text{protein}}^{-1}$, black in A–D). The strains with the integrated P_KAR2(FL)-sfGFP cassette for the monitoring of UPR and producing EcPGA (A), CaLB (C), or TlXynA (D), and the control strain with the integrated x-sfGFP cassette producing EcPGA (B) were cultured under the same cultivation conditions (30°C, pH 5.5, μ_methanol 0.016 h⁻¹).