Fig. 4: The protein payload in OMVs is superior in terms of protein stability and catalytic activity in simulated intestinal fluid.
From: Oral delivery of therapeutic proteins by engineered bacterial type zero secretion system
a Schematic of the Uox secretion approach involving the direct protein secretion apparatus T1SS or modified T0SS-based protein secretion system. Created in BioRender. Gong, X. (2025) https://BioRender.com/r83g400. b Western blot analysis of Uox abundance in the intracellular fraction, supernatant and OMVs of EcN∆nlpI∆thyA-SU and EcN∆thyA-SUT using an anti-His antibody. The stable plasmid system did not affect the function of the recombinant EcN strains outfitted with T0SS or T1SS. c EcN∆nlpI∆thyA-SU with T0SS and EcN∆thyA-SUT with T1SS presented similar UA degradation rates in M9 medium supplemented with 0.5 mM UA under microaerobic culture condition (n = 3 biologically independent samples). d EcN∆nlpI∆thyA-SU with T0SS exhibited greater UA degradation efficiency than that of EcN∆thyA-SUT with T1SS in simulated intestinal fluid supplemented with 0.5 mM UA under microaerobic culture condition (n = 3 biologically independent samples). e Comparison of the enzyme activities of purified protein (T1SS-Uox) or protein-loaded OMVs (OMV-Uox) from EcN∆thyA-SUT or EcN∆nlpI∆thyA-SU, respectively, tested in PBS at various time points at 37 °C (n = 3 biologically independent samples). f, g The stabilities of T1SS-Uox and OMV-Uox were assessed by monitoring the enzyme activities (f) and Uox protein levels (g) in the simulated intestinal fluid at various time points at 37 °C (n = 3 biologically independent samples). The data are presented as the mean ± SD. The P value was determined by two-way analysis of variance (ANOVA) with Tukey’s multiple comparisons test. Source data are provided as a Source Data file.
