Noncovalent immobilization of streptavidin on in vitro- and in vivo-biotinylated bacterial magnetic particles.
Biotinylated magnetic nanoparticles were constructed by displaying biotin acceptor peptide (BAP) or biotin carboxyl carrier protein (BCCP) on the surface of bacterial magnetic particles (BacMPs) synthesized by Magnetospirillum magneticum AMB-1. BAP-displaying BacMPs (BAP-BacMPs) were extracted from bacterial cells and incubated with biotin and Escherichia coli biotin ligase. Then the in vitro biotinylation of BAP-BacMPs was confirmed using alkaline phosphatase-labeled antibiotin antibody. In contrast, BacMPs displaying the intact 149 residues of AMB-1 BCCP (BCCP-BacMPs) and displaying the COOH-terminal 78 residues of BCCP (BCCP78-BacMPs) were biotinylated in AMB-1 cells. The in vivo biotinylation of BCCP-BacMPs and BCCP78-BacMPs was thought to be performed by endogenous AMB-1 biotin ligase. Streptavidin was introduced onto biotinylated BacMPs by simple mixing. In an analysis using tetramethyl rhodamine isocyanate-labeled streptavidin, approximately 15 streptavidin molecules were shown to be immobilized on a single BCCP-BacMP. Furthermore, gold nanoparticle-BacMP composites were constructed via the biotin-streptavidin interaction. The conjugation system developed in this work provides a simple, low-cost method for producing biotin- or streptavidin-labeled magnetic nanoparticles. Various functional materials can be site selectively immobilized on these specially designed BacMPs. By combining the site-selective biotinylation technology and the protein display technology, more innovative and attractive magnetic nanomaterials can be constructed.
Maeda
Yoshiaki
Y
Department of Biotechnology, Tokyo University of Agriculture and Technology, 2-24-16 Naka-cho, Koganei, Tokyo 184-8588, Japan.
Yoshino
Tomoko
T
Takahashi
Masaaki
M
Ginya
Harumi
H
Asahina
Junko
J
Tajima
Hideji
H
Matsunaga
Tadashi
T
D000103:Acetyl-CoA Carboxylase / Q000737:chemistry; D019921:Biotinylation / Q000379:methods*; D019731:Carbon-Nitrogen Ligases / Q000737:chemistry; D002352:Carrier Proteins / Q000737:chemistry; D004926:Escherichia coli / Q000737:chemistry; D029968:Escherichia coli Proteins / Q000737:chemistry; D054889:Fatty Acid Synthase, Type II; D008280:Magnetics; D042201:Magnetospirillum / Q000737:chemistry*; D053758:Nanoparticles / Q000737:chemistry*; D012097:Repressor Proteins / Q000737:chemistry; D019809:Streptavidin / Q000737:chemistry*
D002352:Carrier Proteins; D029968:Escherichia coli Proteins; D012097:Repressor Proteins; D019809:Streptavidin; D054889:Fatty Acid Synthase, Type II; D019731:Carbon-Nitrogen Ligases; C058125:birA protein, E coli; D000103:Acetyl-CoA Carboxylase; C079633:biotin carboxyl carrier protein