In vivo biotinylation of bacterial magnetic particles by a truncated form of escherichia coli biotin ligase and biotin acceptor peptide

Yoshiaki Maeda, Tomoko Yoshino, Tadashi Matsunaga

Research output: Contribution to journalArticle

14 Citations (Scopus)

Abstract

Escherichia coli biotin ligase can attach biotin molecules to a lysine residue of biotin acceptor peptide (BAP), and biotinylation of particular BAP-fused proteins in cells was carried out by coexpression of E. coli biotin ligase (in vivo biotinylation). This in vivo biotinylation technology has been applied for protein purification, analysis of protein localization, and protein-protein interaction in eukaryotic cells, while such studies have not been reported in bacterial cells. In this study, in vivo biotinylation of bacterial magnetic particles (BacMPs) synthesized by Magnetospirillum magneticum AMB-1 was attempted by heterologous expression of E. coli biotin ligase. To biotinylate BacMPs in vivo, BAP was fused to a BacMP surface protein, Mms13, and E. coli biotin ligase was simultaneously expressed in the truncated form lacking the DNA-binding domain. This truncationbased approach permitted the growth of AMB-1 transformants when biotin ligase was heterologously expressed. In vivo biotinylation of BAP on BacMPs was confirmed using an alkaline phosphatase-conjugated antibiotin antibody. The biotinylated BAP-displaying BacMPs were then exposed to streptavidin by simple mixing. The streptavidin-binding capacity of BacMPs biotinylated in vivo was 35-fold greater than that of BacMPs biotinylated in vitro, where BAP-displaying BacMPs purified from bacterial cells were biotinylated by being mixed with E. coli biotin ligase. This study describes not only a simple method to produce biotinylated nanomagnetic particles but also a possible expansion of in vivo biotinylation technology for bacterial investigation.

Original languageEnglish
Pages (from-to)5785-5790
Number of pages6
JournalApplied and Environmental Microbiology
Volume76
Issue number17
DOIs
Publication statusPublished - 2010 Sep
Externally publishedYes

ASJC Scopus subject areas

  • Applied Microbiology and Biotechnology
  • Food Science
  • Biotechnology
  • Ecology

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