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Co-expression of antibody fab heavy and light chain genes from separate evolved compatible replicons in E. coli.

Abstract：

:Antibody molecules bind to antigen with six complementary determining region (CDR) loops, three of which are located on each variable heavy (V(H)) and light (V(L)) chains. Discovery and optimization of antibodies that bind antigen using in vitro techniques require diversification of one or more of these CDRs. Since antibodies are dimeric, simultaneous diversification of heavy and light chains on separate genetic elements would allow "chain shuffling" to occur simply and efficiently. Efficient expression of antibody V(H) and V(L) requires that the two separate replicons be compatible with one another, but also have similar properties, such as copy number in E. coli. Standard plasmids that are compatible with one another in E. coli exist at widely variable copy numbers. Recently we described the isolation of ColE1 mutants that have similar copy numbers but different incompatibility characteristics. Thus, new compatibility groups in the ColE1 family were established. Herein we describe the E. coli expression of V(H) and V(L) genes to form a functional Fab. The ability to express antibody heavy and light chains from separate but compatible high copy plasmids should allow new opportunities in antibody engineering, such as rapid chain shuffling and generation of more complex antibody libraries.

journal_name

J Immunol Methods

journal_title

Journal of immunological methods

authors

Leonard B,Sharma V,Smider V

doi

10.1016/j.jim.2006.09.006

subject

Has Abstract

pub_date

2006-12-20 00:00:00

pages

56-63

issue

1-2

eissn

0022-1759

issn

1872-7905

pii

S0022-1759(06)00260-2

journal_volume

317

pub_type

杂志文章

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