Design of multivalent complexes using the barnase·barstar module

Sergey M. Deyev, Robert Waibel, Ekaterina N. Lebedenko, August P. Schubiger, Andreas Plückthun

Research output: Contribution to journalArticlepeer-review

144 Citations (Scopus)

Abstract

The ribonuclease barnase (12 kDa) and its inhibitor barstar (10 kDa) form a very tight complex in which all N and C termini are accessible for fusion. Here we exploit this system to create modular targeting molecules based on antibody scFv fragment fusions to barnase, to two barnase molecules in series and to barstar. We describe the construction, production and purification of defined dimeric and trimeric complexes. Immobilized barnase fusions are used to capture barstar fusions from crude extracts to yield homogeneous, heterodimeric fusion proteins. These proteins are stable, soluble and resistant to proteolysis. Using fusions with anti-p185HER2-ECD 4D5 scFv, we show that the anticipated gain in avidity from monomer to dimer to trimer is obtained and that favorable tumor targeting properties are achieved. Many permutations of engineered multispecific fusion proteins become accessible with this technology of quasi-covalent heterodimers.

Original languageEnglish
Pages (from-to)1486-1492
Number of pages7
JournalNature Biotechnology
Volume21
Issue number12
DOIs
Publication statusPublished - Dec 2003
Externally publishedYes

ASJC Scopus subject areas

  • Biotechnology
  • Bioengineering
  • Applied Microbiology and Biotechnology
  • Biomedical Engineering
  • Molecular Medicine

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