Production of a recombinant Fab in Pichia pastoris from a Monocistronic expression vector.
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 harbouring a KEX2 processing site between both chains. After KEX cleavage, a correctly folded mature Fab is formed. The produced recombinant protein is characterized as a heterodimeric functional Fab. The vector described is a new tool for the proper expression of antibody fragments or any heterodimeric polypeptides.
Universidade de Brasília, 70910-900, Brasília, DF, Brazil.
Rafael Trindade
RT
Burtet
Marcos Antônio
MA
Santos-Silva
Guilherme Antônio Marques
GA
Buss
Lidia Maria Pepe
LM
Moraes
Andrea Queiroz
AQ
Maranhão
Marcelo Macedo
MM
Brigido
D019281:Dimerization; D015870:Gene Expression; D005822:Genetic Vectors; D007140:Immunoglobulin Fab Fragments / Q000096:biosynthesis* / Q000235:genetics / Q000276:immunology; D007700:Kinetics; D010843:Pichia / Q000235:genetics*; D011994:Recombinant Proteins / Q000096:biosynthesis* / Q000737:chemistry / Q000276:immunology
D007140:Immunoglobulin Fab Fragments; D011994:Recombinant Proteins