Evaluation of three different vectors on human iLRP expression in Escherichia coli

Abstract

Background: iLRP (immature laminin receptor protein) is a tumor associated antigen over-expressed on the surface of most human cancer cells and plays an important role in the process of tumorigenesis and development. It has strong auto-immunogenicity in patients and is a good target protein for tumor immunotherapy. The aim of this study is to find a practical and an efficient method for the production of recombinant iLRP, and to provide enough recombinant protein for further study of its potential applications.Results: In this report, three different expression vectors based on pET-30a(+) from pET expression systems were constructed. The first one is to add a 6xHis-Tag to the N-terminal of natural iLRP, which is called as pET-His-iLRP. The second one is to include an S-Tag between the C-terminal of 6xHis-Tag and the N-terminal of natural iLRP, which is called as pET-His-S-iLRP. The third one only contains the 6xHis-Tag in front of the N-terminal of iLRP, but the natural iLRP gene was first optimized according to the bacterial genome, and the constructed vector was named as pET-His-Opt-iLRP. Then the expression of three vectors in Escherichia coli (E. coli) BL21 (DE3) was analyzed. Results demonstrated that the expression of iLRP was the highest in the vector of pET-His-Opt-iLRP, followed by the vector of pET-His-S-iLRP, and the vector of pET-His-iLRP expressed the lowest iLRP.Conclusions: According to the results, it was concluded that the vetor pET-His-Opt-iLRP combining the codon-optimized iLRP gene with a 6xHis-Tag had the best effect on the expression of iLRP, and the expression in vectors with the natural iLRP gene depends on its leader sequences in the expression frame. The longer leader sequences accommodate to the host E. coli, the higher expression the engineered gene would be.