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Open Access Research

In vivo modification of tyrosine residues in recombinant mussel adhesive protein by tyrosinase co-expression in Escherichia coli

Yoo Seong Choi1, Yun Jung Yang2, Byeongseon Yang2 and Hyung Joon Cha23*

Author Affiliations

1 Department of Chemical Engineering, Chungnam National University, Daejon, 305-764, Korea

2 Department of Chemical Engineering, Pohang University of Science and Technology, Pohang, 790-784, Korea

3 Ocean Science and Technology Institute, Pohang University of Science and Technology, Pohang, 790-784, Korea

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Microbial Cell Factories 2012, 11:139  doi:10.1186/1475-2859-11-139

Published: 24 October 2012

Abstract

Background

In nature, mussel adhesive proteins (MAPs) show remarkable adhesive properties, biocompatibility, and biodegradability. Thus, they have been considered promising adhesive biomaterials for various biomedical and industrial applications. However, limited production of natural MAPs has hampered their practical applications. Recombinant production in bacterial cells could be one alternative to obtain useable amounts of MAPs, although additional post-translational modification of tyrosine residues into 3,4-dihydroxyphenyl-alanine (Dopa) and Dopaquinone is required. The superior properties of MAPs are mainly attributed to the introduction of quinone-derived intermolecular cross-links. To solve this problem, we utilized a co-expression strategy of recombinant MAP and tyrosinase in Escherichia coli to successfully modify tyrosine residues in vivo.

Results

A recombinant hybrid MAP, fp-151, was used as a target for in vivo modification, and a dual vector system of pET and pACYC-Duet provided co-expression of fp-151 and tyrosinase. As a result, fp-151 was over-expressed and mainly obtained from the soluble fraction in the co-expression system. Without tyrosinase co-expression, fp-151 was over-expressed in an insoluble form in inclusion bodies. The modification of tyrosine residues in the soluble-expressed fp-151 was clearly observed from nitroblue tetrazolium staining and liquid-chromatography-mass/mass spectrometry analyses. The purified, in vivo modified, fp-151 from the co-expression system showed approximately 4-fold higher bulk-scale adhesive strength compared to in vitro tyrosinase-treated fp-151.

Conclusion

Here, we reported a co-expression system to obtain in vivo modified MAP; additional in vitro tyrosinase modification was not needed to obtain adhesive properties and the in vivo modified MAP showed superior adhesive strength compared to in vitro modified protein. It is expected that this co-expression strategy will accelerate the use of functional MAPs in practical applications and can be successfully applied to prepare other Dopa/Dopaquinone-based biomaterials.

Keywords:
Mussel adhesive protein; Dopa; Dopaquinone; In vivo modification; Tyrosinase; Co-expression; Escherichia coli