The browser you are using is not supported by this website. All versions of Internet Explorer are no longer supported, either by us or Microsoft (read more here: https://www.microsoft.com/en-us/microsoft-365/windows/end-of-ie-support).

Please use a modern browser to fully experience our website, such as the newest versions of Edge, Chrome, Firefox or Safari etc.

Carl Borrebaeck

Carl Borrebaeck

Professor

Carl Borrebaeck

A point mutation in a murine immunoglobulin V-region strongly influences the antibody yield in Escherichia coli

Author

  • M. Dueñas
  • M. Ayala
  • J. Vázquez
  • M. Ohlin
  • E. Söderlind
  • C. A K Borrebaeck
  • J. V. Gavilondo

Summary, in English

Recombinant DNA technology has made it possible to produce specific Fab and scFv antibody (Ab) fragments in prokaryotic host cells. Using vectors designed for periplasmic expression of encoded Ab fragments, we have been studying how the sequence and genetic localization of the light chain (L-chain) variable region gene of a mouse Ab (CB-Nm.1) determined the level of Ab production. The variable region was shown to belong to the VKV family and contained a previously unreported Ile72. Nine different Ab constructions were tested in monocistronic (scFv) or dicistronic (Fab) operons for their ability to affect the synthesis level of the L-chain. When the gene coding for the L-chain was located downstream from the Fd fragment gene, the substitution of codons encoding Ile by a codon encoding Thr was found to be crucial for any expression of the L-chain fragment. This was, however, not accompanied by an increase in L-chain-specific mRNA, neither was there any change in the size of the mRNA. The fact that the unmutated L-chain protein was produced from cells transformed with certain other constructions indicated that the protein as such was not incompatible with the prokaryotic environment. Together, this suggested that the translation process was involved in the restricted production of the L-chain. Thus, surprisingly small substitutions significantly affected the expression level, a fact that will have important implications on the library size expressed in prokaryotic hosts, including phagedisplayed Ab libraries.

Department/s

  • Department of Immunotechnology

Publishing year

1995

Language

English

Pages

61-66

Publication/Series

Gene

Volume

158

Issue

1

Document type

Journal article

Publisher

Elsevier

Keywords

  • Antibody fragment
  • E. coli expression
  • Fab
  • phage display vector
  • scFv

Status

Published

ISBN/ISSN/Other

  • ISSN: 0378-1119