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ABSTRACTThe emulsifying ability of SA01-OmpA (outer membrane protein A from Acinetobacter sp. SA01) was found to be constrained by challenges like low production efficiency and high costs associated with protein recovery from E. coli inclusion bodies, as described in our previous study. The present study sought to benefit from the advantages of the targeted truncating of SA01-OmpA protein, taking into account the reduced propensity of protein expression as inclusion bodies and cytotoxicity. Here, the structure and activity relationship of two truncated recombinant forms of SA01-OmpA protein was unraveled through a hybrid approach based on experimental data and computational methodologies, representing an innovative bioemulsifier with advantageous emulsifying activity. The recombinant truncated SA01-OmpA variants were cloned and heterologously expressed in E. coli host cells and subsequently purified. The results showed increased emulsifying activity of N-terminally truncated SA01-OmpA (NT-OmpA) compared to full-length SA01-OmpA. Molecular dynamics (MD) simulations analysis demonstrated a direct correlation between the C-terminally truncated SA01-OmpA (CT-OmpA) and its expression as inclusion bodies. Analysis of the structure-activity relationship of truncated variants of SA01-OmpA revealed that, compared to the full-length protein, deletion of the β-barrel portion from the N-terminal of SA01-OmpA increased the emulsifying activity of NT-OmpA while lowering its expression as inclusion bodies. Contrary to the full-length protein, the N-terminally truncated SA01-OmpA was not as cytotoxic, according to the MTT assay, FCM analysis, and AO/EB staining. The findings of this extensive study advance our knowledge of SA01-OmpA at the molecular level as well as the design and development of efficient bioemulsifiers.IMPORTANCEPrevious research (Shahryari et al. 2021, mSystems 6: e01175-20) introduced and characterized the SA01-OmpA protein as a multifaceted protein with a variety of functions, including maintaining cellular homeostasis under oxidative stress conditions, biofilm formation, outer membrane vesicles (OMV) biogenesis, and beneficial emulsifying capacity. By truncating the SA01-OmpA protein, the current study presents a unique method for developing protein-type bioemulsifiers. The findings indicate that the N-terminally truncated SA01-OmpA (NT-OmpA) has the potential to fully replace full-length SA01-OmpA as a novel bioemulsifier with significant emulsifying activity. This study opens up a new frontier in bioemulsifiers, shedding light on a possible relationship between the structure and activity of SA01-OmpA truncated forms.

BackgroundDrug resistance in cancer cells is a major concern in chemotherapy. Cisplatin (CIS) is one of the most effective chemotherapeutics for ovarian cancer. Here, we investigated an experimental approach to increase CIS cytotoxicity and overcome cell resistance using nanoparticle-based combination treatments.MethodsPolyethylenimine (PEI)-based magnetic iron oxide nanocomplexes were used for drug delivery in genetically matched CIS-resistant (A2780/CP) and -sensitive (A2780) ovarian cancer cells in the presence of a 20 mT static magnetic field. Magnetic nanoparticles (MNPs) were synthesized and bonded to PEI cationic polymers to form binary complexes (PM). The binding of CIS to the PM binary complexes resulted in the formation of ternary complexes PM/C (PEI–MNP/CIS) and PMC (PEI–MNP–CIS).ResultsCIS cytotoxicity increased at different concentrations of CIS and PEI in all binary and ternary delivery systems over time. Additionally, CIS induced cell cycle arrest in the S and G2/M phases and reactive oxygen species production in both cell lines. Ternary complexes were more effective than binary complexes at promoting apoptosis in the treated cells.ConclusionPEI-based magnetic nanocomplexes can be considered novel carriers for increasing CIS cytotoxicity and likely overcoming drug resistance of ovarian cancer cells.