Enzymatic labeling of a single chain variable fragment of an antibody with alkaline phosphatase by microbial transglutaminase

Takeshi Takazawa, Noriho Kamiya, Hiroshi Ueda, Teruyuki Nagamune

Research output: Contribution to journalArticle

35 Citations (Scopus)

Abstract

Functional cross-linking of a single chain Fv fragment of anti-hen egg-white lysozyme antibody (scFv) and alkaline phosphatase (AP) was explored using microbial transglutaminase (MTG) from Streptomyces mobaraensis. A specific peptidyl linker for MTG was genetically fused to the N-terminus of each protein and the resultant proteins were obtained separately by bacterial expression. The recombinant peptide-tagged scFv and AP were site-specifically cross-linked by MTG through the extra peptidyl linkers in vitro, which mainly yielded the heterodimer (i.e., scFv-AP conjugate). The enzymatic cross-linking reaction had little influence on either the antigen-binding ability of the scFv moiety or the enzymatic activity of the AP moiety of the conjugate, allowing use within an enzyme-linked immunosorbent assay. The results obtained suggest that the enzymatic approach with MTG facilitates the posttranslational construction of functional fusion proteins.

Original languageEnglish
Pages (from-to)399-404
Number of pages6
JournalBiotechnology and Bioengineering
Volume86
Issue number4
DOIs
Publication statusPublished - May 20 2004

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Single-Chain Antibodies
Transglutaminases
Phosphatases
Antibodies
Labeling
Alkaline Phosphatase
Proteins
Enzymes
Immunoglobulin Variable Region
Egg White
Immunosorbents
Cross Reactions
Streptomyces
Antigens
Muramidase
Peptides
Assays
Fusion reactions
Enzyme-Linked Immunosorbent Assay

All Science Journal Classification (ASJC) codes

  • Biotechnology
  • Bioengineering
  • Applied Microbiology and Biotechnology

Cite this

Enzymatic labeling of a single chain variable fragment of an antibody with alkaline phosphatase by microbial transglutaminase. / Takazawa, Takeshi; Kamiya, Noriho; Ueda, Hiroshi; Nagamune, Teruyuki.

In: Biotechnology and Bioengineering, Vol. 86, No. 4, 20.05.2004, p. 399-404.

Research output: Contribution to journalArticle

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