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Journal of Biochemistry Advance Access first published online on November 24, 2007
This version published online on November 27, 2007
Journal of Biochemistry, doi:10.1093/jb/mvm226
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© 2007 The Japanese Biochemical Society

Rapid Communication

Production of a recombinant Fab in Pichia pastoris from a monocistronic expression vector

Rafael Trindade Burtet1,2, Marcos Antônio Santos-Silva1, Guilherme Antônio Marques Buss1, Lidia Maria Pepe Moraes1, Andrea Queiroz Maranhão1,2 and Marcelo Macedo Brigido1,2,*

1Universidade de Brasília, 70910-900, Brasília, DF, Brazil
2Instituto de Investigação em Imunologia

*Corresponding author: Dr. Marcelo Macedo Brigido, Departamento de Biologia Celular-IB, Universidade de Brasília, 70.910-900 Brasília, DF, Brasil,, Tel. 55 (61) 3072423; Fax. 55 (61) 3498411; E-mail: brigido{at}unb.br

Received October 16, 2007; Accepted November 15, 2007


  Abstract

Recombinant Fab is usually expressed using dicistronic vectors producing the heavy and light chains separately. We developed an improved vector for Fab fragment expression in Pichia pastoris, which allows a stoichiometric expression of both chains based on a monocistronic arrangement. The protein is produced as a unique polypeptide harboring a KEX2 processing site between both chains. After KEX cleavage, a correctly folded mature Fab is formed. The produced recombinant protein was characterized as a heterodimeric functional Fab.The vector described is a new tool for the proper expression of antibody fragments or any heterodimeric polypeptides.

Key Words: Antibody engineering, Pichia pastoris, Fab expression, Kex


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