Soo Youn Lee

Inorganic nanomaterial-based biocatalysts.

Over the years, nanostructures have been developed to enable to support enzyme usability to obtain highly selective and efficient biocatalysts for catalyzing processes under various conditions. This review summarizes recent developments in the nanostructures for enzyme supporters, typically those formed with various inorganic materials. To improve enzyme attachment, the surface of nanomaterials is properly modified to express specific functional groups. Various materials and nanostructures can be applied to improve both enzyme activity and stability. The merits of the incorporation of enzymes in inorganic nanomaterials and unprecedented opportunities for enhanced enzyme properties are discussed. Finally, the limitations encountered with nanomaterial-based enzyme immobilization are discussed together with the future prospects of such systems.

High throughput detection and selective enrichment of histidine-tagged enzymes with Ni-doped magnetic mesoporous silica

This work reports a simple and facile method to prepare novel magnetic mesoporous silica (MMS) materials with high magnetic strength for the convenient and high throughput detection of histidine-tagged enzymes with Ni-doped surfaces. These materials are designed by the incorporation of high-abundance and homogeneously dispersed iron nanoparticles within the mesopores by thermal hydrogen reduction after the incorporation of ferrous ions and demonstrated the selective enrichment and high-throughput recognition of His-tagged enzymes with multi-point anchoring by selective conjugation between the His-tag and Ni ions. Selective His-tagged enzyme enrichment efficiency was compared with nickel-based MMS materials, such as Ni2+-MMS and Ni-MMS, and nickel ion doped silica-coated magnetic nanoparticles (Ni2+-MNPs). The efficiency was calculated to be 100 ± 1.93%, 70.94 ± 1.95%, and 37.03 ± 5.93% for Ni2+-MMS, Ni-MMS, and Ni2+-MNPs, respectively. This method enables a high-throughput and advanced systematic approach for the separation and immobilization of proteins which cover a broad spectrum of polyhistidine-tagged proteins.

Amino acid side chain-like surface modification on magnetic nanoparticles for highly efficient separation of mixed proteins

This work reports on the realization of specific functionalized silica-coated magnetic nanoparticles (Si-MNPs) for effective protein separation through surface modification with various amino acid side chain-like functional groups such as thiol (-SH), disulfide (-S-S-), carbon chain (-C(n)), carboxyl (-COOH), amine (-NH(2)), and aldehyde (-CHO). This study also suggests an improved and convenient method for the synthesis of functionalized Si-MNPs by hydrolysis condensation with silan-coupling agents. The protein adsorption effects in a coexistent mixed state are explored using various proteins, which have different isoelectric point (pI) values and molecular weights, in order to elucidate the binding performance of different proteins one solution. The adsorption efficiency of bovine serum albumin (BSA; 66 kDa; pI=4.65) and lysozyme (LYZ; 14.3 kDa; pI=11) is 70-100% with various amino acid side chain-like functional groups. However, the adsorption efficiency of a mixed protein solution of BSA and LYZ was different. Although the relatively bulky BSA molecule displayed 50% and 20% adsorption corresponding to pH 4.65, and pH 11, respectively, the smaller LYZ provided almost 100% adsorption at both pH 4.65 and pH 11.

Enhancing immunoassay detection of antigens with multimeric protein Gs

This paper describes a method for the effective and self-oriented immobilization of antibodies on magnetic silica-nanoparticles using a multimeric protein G. Cysteine-tagged recombinant dimers and trimers of protein G were produced in Escherichia coli BL21 by repeated linking of protein G monomers with a flexible (GGGGS)(3) linker. Amino-functionalized silica-coated magnetic nanoparticles (SiO(2)-MNPs, Fe(3)O(4)@SiO(2)) were prepared and coupled to the protein G multimers, giving the final magnetic immunosensor. The optimal conditions for the reaction between the protein Gs and the SiO(2)-MNPs was a time of 60 min and a concentration of 100 μg/mL, resulting in coupling efficiencies of 77%, 67% and 55% for the monomeric, dimeric and trimeric protein Gs, respectively. Subsequently, anti-hepatitis B surface antigen (HBsAg) was immobilized onto protein G-coupled SiO(2)-MNPs. The quantitative efficiency of antibody immobilization found the trimeric protein G to be the best, followed by the dimeric and monomeric proteins, which differs from the coupling efficiencies. Using all three protein constructs in an HBsAg fluoroimmunoassay, the lowest detectable concentrations were 500, 250 and 50 ng/mL for the monomeric, dimeric and trimeric protein G-coupled SiO(2)-MNPs, respectively. Therefore, multimeric protein Gs, particularly the trimeric form, can be employed to improve antibody immobilization and, ultimately, enhance the sensitivity of immunoassays. In addition, the multimeric protein Gs devised in this study can be utilized in other immunosensors to bind the antibodies at a high efficiency and in the proper orientation.

Rapid pathogen detection with bacterial-assembled magnetic mesoporous silica

We report rapid and accurate pathogen detection by coupling with high efficiency magnetic separation of pathogen by Ni(2+)-heterogeneous magnetic mesoporous silica (Ni-HMMS) and real time-polymerase chain reaction (RT-PCR) technique. Ni-HMMS was developed with a significant incorporation of Fe particles within the silica mesopores by programmed thermal hydrogen reaction and functionalized with Ni(2+) ion on the surface by the wet impregnation process. High abundant Ni(2+) ions on the Ni-HMMS surface were able to assemble with cell wall component protein NikA (nickel-binding membrane protein), which contains several pathogenic bacteria including Escherichia coli O157:H7. NikA protein expression experiment showed the outstanding separation rate of the nikA gene-overexpressed E. coli (pSY-Nik) when comparing with wild-type E. coli (44.5 ± 13%) or not over-expressed E. coli (pSY-Nik) (53.2 ± 2.7%). Moreover, Ni-HMMS showed lower obstacle effect by large reaction volume (10 mL) than spherical core/shell-type silica magnetic nanoparticles functionalized with Ni(2+) (ca. 40 nm-diameters). Finally, the Ni-HMMS was successfully assessed to separate pathogenic E. coli O157:H7 and applied to direct and rapid RT-PCR to quantitative detection at ultralow concentration (1 Log10 cfu mL(-1)) in the real samples (milk and Staphylococcus aureus culture broth) without bacterial amplification and DNA extraction step.

Antimicrobial medical sutures with caffeic acid phenethyl ester and their in vitro/in vivo biological assessment

The work report the in vitro and in vivo assessment of antimicrobial poly(lactic-co-glycolic acid) (PLGA) sutures loaded with noble natural extracts, caffeic acid phenethyl ester (CAPE) from natural propolis. The mechanical characteristics of the medical sutures have been elucidated including tensile strength as a function of biodegradability, and the sustained release profile of CAPE on sutures has been measured as a function of time, concentration and optimization with the aim of maximizing their antimicrobial effect against Staphylococcus aureus and Escherichia DH5α bacteria. Furthermore, in vitro and in vivo antimicrobial assessments – cytotoxicity tests, bacterial reverse mutation (Ames) assays, and micronucleus assays – were achieved to investigate biocompatibility. Real time RT-PCR showed that the antimicrobial effect of CAPE is related to outer membrane damage and reactive oxygen species synthesis in bacteria.

Rapid and selective separation for mixed proteins with thiol functionalized magnetic nanoparticles

Thiol group functionalized silica-coated magnetic nanoparticles (Si-MNPs@SH) were synthesized for rapid and selective magnetic field-based separation of mixed proteins. The highest adsorption efficiencies of binary proteins, bovine serum albumin (BSA; 66 kDa; pI = 4.65) and lysozyme (LYZ; 14.3 kDa; pI = 11) were shown at the pH values corresponding to their own pI in the single-component protein. In the mixed protein, however, the adsorption performance of BSA and LYZ by Si-MNPs@SH was governed not only by pH but also by the molecular weight of each protein in the mixed protein.

Histidine-tagged enzyme conjugated heterogeneous magnetic mesoporous silica for high efficient biodegradation of catechol

In this study, we present a novel method for mesoporous silicas which are functionalized with Fe and Ni ion to support His-tagged proteins. Mesoporous silicas were combined with iron (II) chloride tetrahydrate for initiating magnetizing process, followed by reduction in H2 media. The nickel (II) chloride hexahydrate was introduced into pore and surface of heterogeneous magnetic mesoporous silicas (HMMS) by wet impregnation process. The resulting materials were reduced in a flowing of H2 at 500 °C for 2h, thereby producing Ni particles on the surface of the HMMS. The Ni-HMMS was characterized by various state-of-the-art techniques, such as WAXS, FE-SEM, TEM, SQUID and nitrogen sorption. Ni-HMMS showed high adsorption level of His-tagged protein from Escherichia coli lysate. In addition, the purified His6-CatA was observed high enzyme activity. It suggests that Ni-MMS system provides contribution for maintain stable protein structure as well as simplified protein purification process.