A novel recombinant chimeric bio-adhesive protein consisting of mussel foot protein 3, 5, gas vesicle protein A, and CsgA curli protein expressed in Pichia pastoris

Nazanin Bolghari,Hosein Shahsavarani,Hadi Habibollahi,Masoumeh Anvari

doi:10.1186/s13568-022-01362-5

Nazanin Bolghari, Hosein Shahsavarani + Show 2 more

Open Access

https://doi.org/10.1186/s13568-022-01362-5

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Abstract

Despite various efforts to produce potent recombinant bio-adhesive proteins for medical purposes, efficient production of a safe and feasible bio-glue is not yet a commercial reality due to the weak properties or low expression levels. Here, a feasible expression system has been developed to produce strong recombinant fusion bioinspired protein using mussel foot protein 3 and 5 (Mfps) along with gas vesicle protein A (GvpA) of Anabaena flos-aquae, and a curli protein CsgA from E. coli, expressed under the control of alcohol oxidase (AOX1) promoter for high-level production in yeast P. pastoris using pPICZα vector. Purified chimeric proteins were first evaluated using western blotting, and their remaining dihydroxyphenylalanine (DOPA) was measured in the modified proteins by NBT assay. We further elucidated the mechanistic properties of obtained adhesive protein assembly in various pH levels based on its different subunits using atomic force microscopy (AFM) when adsorbed onto the mica surface. We found that both combinational structural features of subunits and post-translational changes during expression in yeast host have led to potent adherence due to higher DOPA residues specially in acidic condition and tetrad complex which is higher than that of earlier reports in prokaryotic systems. We believe that our obtained chimeric protein resulted from the fusion of GvpA and CsgA proteins with DOPA-containing Mfp proteins, expressed in the methylotrophic yeast, P. pastoris, not only presents a candidate for future biomedical applications but also provides novel biological clues used for high-performance bioinspired biomaterial designation.Graphical

Highlights

The use of adhesive material for the repairment of tissues following surgery or trauma has attracted much attention and is likely to replace traditional methods such as suture, wiring or staples
Genomic Integration of the pPICZα vector MPF5-CsgA and MPF3-gas vesicle protein A (GvpA) genes were cloned into the yeast cell of Pichia pastoris and produced the desired proteins
Similar from those produced in E. coli, recombinant proteins were expressed as inclusion bodies in yeast Pichia pastoris

Summary

Introduction

The use of adhesive material for the repairment of tissues following surgery or trauma has attracted much attention and is likely to replace traditional methods such as suture, wiring or staples. Most of the surgical adhesives used in operations are cyanoacrylate derivatives, which still have numerous limitations like cytotoxicity, inflammatory reaction or operational complexities (Choi et al 2011). Another type of wound closure generally applied in surgery is animal-derived glues, which have numerous issues such as, safety concerns or non-adequate bond toughness. Bioinspired recombinant bio-adhesive recombinant protein production has been suggested to address the aforementioned challenges with less drawbacks such as safety issues, need for further cosmetic operation or any allergic reactions (Cha et al 2009). Recent advances in biomimetic research have recently led to the design of various novel recombinant proteinaceous glues capable of satisfying specific demands of both biomedical and technical fields through cloning and expression of the natural adhesive proteins (Deming 1999)

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