Methodology article

Evolutional selection of a combinatorial phage library displaying randomly-rearranged various single domains of immunoglobulin (Ig)-binding proteins (IBPs) with four kinds of Ig molecules

Hua Yang^1* , Jie Cao^2* , Lian-Qing Li³ , Xia Zhou¹ , Qiu-Li Chen² , Wen-Ting Liao² , Zong-Mei Wen² , Shao-Hua Jiang² , Rong Xu² , Jian-An Jia² , Xin Pan² , Zhong-Tian Qi² and Wei Pan²

¹  Department of Microbiology and Immunology, Shan Xi Medical University, Tai-yuan, 030001, PR China

²  Department of Microbiology, State Key Laboratory of Medical Immunology, Second Military Medical University, Shanghai 200433, PR China

³  Center of clinical diagnosis, Shan Xi province, Tai-yuan, 030012, PR China

author email corresponding author email* Contributed equally

BMC Microbiology 2008, 8:137doi:10.1186/1471-2180-8-137

Published:	13 August 2008

Abstract

Background

Protein A, protein G and protein L are three well-defined immunoglobulin (Ig)-binding proteins (IBPs), which show affinity for specific sites on Ig of mammalian hosts. Although the precise functions of these molecules are not fully understood, it is thought that they play an important role in pathogenicity of bacteria. The single domains of protein A, protein G and protein L were all demonstrated to have function to bind to Ig. Whether combinations of Ig-binding domains of various IBPs could exhibit useful novel binding is interesting.

Results

We used a combinatorial phage library which displayed randomly-rearranged various-peptide-linked molecules of D and A domains of protein A, designated PA(D) and PA(A) respectively, B2 domain of protein G (PG) and B3 domain of protein L (PL) for affinity selection with human IgG (hIgG), human IgM (hIgM), human IgA (hIgA) and recombinant hIgG1-Fc as bait respectively. Two kinds of novel combinatorial molecules with characteristic structure of PA(A)-PG and PA(A)-PL were obtained in hIgG (hIgG1-Fc) and hIgM (hIgA) post-selection populations respectively. In addition, the linking peptides among all PA(A)-PG and PA(A)-PL structures was strongly selected, and showed interestingly divergent and convergent distribution. The phage binding assays and competitive inhibition experiments demonstrated that PA(A)-PG and PA(A)-PL combinations possess comparable binding advantages with hIgG/hIgG1-Fc and hIgM/hIgA respectively.

Conclusion

In this work, a combinatorial phage library displaying Ig-binding domains of protein A, protein G, or protein L joined by various random linking peptides was used to conducted evolutional selection in vitro with four kinds of Ig molecules. Two kinds of novel combinations of Ig-binding domains, PA(A)-PG and PA(A)-PL, were obtained, and demonstrate the novel Ig binding properties.