Surface modification is an important part of fabricating nanoparticles with specific properties and functions. We have designed a dipeptide, which we call NS polypeptide, that consists of four asparagine (N) residues and one serine (S) residue, as a molecule for nanoparticle surface modification. Surface modification of magnetic nanoparticles with the NS polypeptide results in reduction of particle-particle and particle-cell interactions. Here, we describe the surface modification and functionalization of bacterial magnetic particles (BacMPs) by spontaneous integration of temporin L conjugated to NS polypeptide. BacMP membranes were modified temporin L. Furthermore, peptide-modified BacMPs showed high dispersibility in aqueous solution compared to unmodified BacMPs. This surface modification technique may represent a new strategy for reducing non-specific binding of nanoparticles to proteins or cells for use in a variety of protein- or cell-associated applications.

Mms6 is a small acidic protein which is tightly bound to magnetite in the bacterium Magnetospirillum magneticum AMB-1. Mms6 has been previously shown to promote iron-binding capacity as well as modulate the size and morphology of magnetic iron oxide crystals in vitro. In this study, we synthesized iron oxide crystals by using a monolayer-modified substrate. A self-assembled monolayer of octadecyltrimethoxysilane was modified on a silicon substrate. Recombinant Mms6 protein was attached to the substrate through the hydrophobic interactions between the protein molecules and the monolayer. The immobilization of protein molecules on the substrate surface was confirmed by fluorescent labeling of these molecules and subsequent fluorescence microscopy. This protein-modified substrate was then used as a template for iron oxide crystal formation in a ferrous solution. Scanning electron microscopy revealed site-specific formation of iron oxide crystals in substrate regions with immobilized proteins. This use of proteins might provide an alternative method for the bottom-up fabrication of nano-sized magnetic particles.

Magnetotactic bacteria are a unique species of bacteria, commonly recognized by the presence of magnetic particles within them. These intracellular, nanosized magnetic particles enable the bacteria to migrate and be manipulated by magnetic force. To date, magnetotactic bacteria have been widely researched and implemented in various biotechnology based applications. In this study, as an extension to its applications, the magnetotactic bacterium, Magnetospirillum magneticum AMB-1, was applied in the microbial recovery of gold from plating waste. M. magneticum AMB-1 successfully precipitated approximately 42% and 100% of gold from growth medium containing 10 μM gold and from a mixture of plating waste/growth medium containing 0.4 μM gold, respectively. These observations and results strongly suggests that an important advancement in biorecovery of rare metals and bioremediation of toxic metals was achieved in which the application of whole cell bacteria, and direct precipitation of metals from plating waste using magnetotactic bacteria was performed for the first time.

Magnetic separation of target cells from mixtures, such as peripheral blood and bone marrow, has considerable practical potential in research and medical applications. Among the current cell separation techniques, magnetic cell separation using immunomagnetic particles has been routinely applied and has proven rapidness and simplicity. Magnetospirillum magneticum AMB-1 synthesizes intracellular nano-sized bacterial magnetic particles (BacMPs) that are individually enveloped by a stable lipid bilayer membrane. BacMPs, which exhibit strong ferrimagnetism, can be collected easily with commercially available permanent magnets. In this study, a novel magnetic nanoparticle displaying protein G (protein G-BacMPs) was fabricated, and one-step cell separation for direct cell separation from whole blood was performed using the protein G-BacMPs. B lymphocytes (CD20+ cells), which cover less than 0.3×10−2 % of whole blood cells, were separated with 93% purity using protein G-BacMPs binding with anti-CD20 monoclonal antibodies. The results of this study demonstrate the utility of protein G-BacMPs and the magnetic cell separation approach based on protein G-BacMPs in numerous applications.

Biotinylated magnetic nanoparticles were constructed by displaying biotin acceptor peptide (BAP) on the surfaces of bacterial magnetic particles (BacMPs) synthesized by Magnetospirillum magneticum AMB-1. Both BAP and green fluorescent protein (GFP) were fused to Mms13 that was isolated from BacMP membranes. The localization of the fusion protein, BAP-Mms13-GFP, was confirmed by fluorescence analysis. BacMPs that expressed BAP-Mms13-GFP (BAP/GFP-BacMPs) were extracted from bacterial cells and incubated with biotin and Escherichia coli biotin ligase. The in vitro biotinylation of BAP/GFP-BacMPs was confirmed using alkaline phosphatase (ALP)-streptavidin detection. The conjugation system developed in this study provides a method for producing biotin- or streptavidin-labeled magnetic nanoparticles without the use of a crosslinker reagent. Various functional materials can be immobilized site-selectively onto these uniquely designed BacMPs. By combining this site-selective biotinylation technology and protein display methodology, increasingly innovative and attractive magnetic nano-materials can be constructed.

Magnetic particles are used for various biomedical applications because they are easy to both handle and separate from biological samples. Nano-sized bacterial magnetic particles (BacMPs) that display the human estrogen receptor ligand binding domain (ERLBD) on their surfaces were successfully produced by the magnetotactic bacterium, Magnetospirillum magneticum AMB-1. A receptor assay for endocrine-disrupting chemicals using ERLBD-displaying BacMPs was developed. A BacMP membrane-specific protein, Mms16 or Mms13, was used as an anchor protein to localize the ERLBD on the surfaces of BacMPs. ERLBD-BacMP complexes were assayed for competitive binding of alkaline phosphatase-conjugated 17β-estradiol (ALP-E2). Inhibition curve of ALP-E2 to the powerful antagonist, tamoxifen was generated by measuring decreases in luminescence intensity that resulted from the enzymatic reaction of alkaline phosphatase. The overall simplicity of this receptor-binding assay results in a method that can be easily adapted to a high-throughput format.

Nano-technology has been identified as an area which will bring about new evolutions in materials, devices and processes. The challenges of nano-biotechnology entail manufacturing more sophisticated and highly efficient biosensors and biomaterials at the nano-scale level for use in interdisciplinary fields. Here, we introduce a biomaterial produced by magnetic bacteria, bacterial magnetic particles (BMPs), and the molecular architecture technique we have used for its application.

Magnetic bacteria synthesize intracellular magnets which are encapsulated by lipid bilayer membranes. Sizes of BMPs vary from 50 - 100 nm in diameter, and number over 10 per cell. BMPs are composed of magnetite (Fe3O4) with a single magnetic domain. Easy aqueous dispersion of BMPs enable development of highly sensitive chemiluminescence enzyme immunoassays by the chemical coupling of antibodies on BMP surfaces. BMPs can likewise be used as drug delivery systems employing magnetoliposomes with high capture volumes. We previously reported a technique for preparing recombinant BMPs on which proteins were displayed by gene-fusion. We furthermore applied such recombinant BMPs to biotechnologically-important issues, including novel bioassay platforms for medicine and environmental management. We envisage the production of more refined chemicals and agents through expression on BMPs by gene-fusion followed by simple purification using magnet.