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Biomedical subjects

Hyun Gyu Park

Publications and source records attributed to Hyun Gyu Park.

10 recordsLinked to original sources
PubMed2007

Crosslinked enzyme aggregates in hierarchically-ordered mesoporous silica: a simple and effective method for enzyme stabilization.

alpha-chymotrypsin (CT) and lipase (LP) were immobilized in hierarchically-ordered mesocellular mesoporous silica (HMMS) in a simple but effective way for the enzyme stabilization, which was achieved by the enzyme adsorption followed by glutaraldehyde (GA) crosslinking. This resulted in the formation of nanometer scale crosslinked enzyme aggregates (CLEAs) entrapped in the mesocellular pores of HMMS (37 nm), which did not leach out of HMMS through narrow mesoporous channels (13 nm). CLEA of alpha-chymotrypsin (CLEA-CT) in HMMS showed a high enzyme loading capacity and significantly increased enzyme stability. No activity decrease of CLEA-CT was observed for 2 weeks under even rigorously shaking condition, while adsorbed CT in HMMS and free CT showed a rapid inactivation due to the enzyme leaching and presumably autolysis, respectively. With the CLEA-CT in HMMS, however, there was no tryptic digestion observed suggesting that the CLEA-CT is not susceptible to autolysis. Moreover, CLEA of lipase (CLEA-LP) in HMMS retained 30% specific activity of free lipase with greatly enhanced stability. This work demonstrates that HMMS can be efficiently employed as host materials for enzyme immobilization leading to highly enhanced stability of the immobilized enzymes with high enzyme loading and activity.

Chymotrypsin↗
PubMed2006

Poly(dimethyl siloxane)-based protein chip for simultaneous detection of multiple samples: use of glycidyl methacrylate photopolymer for site-specific protein immobilization.

This paper describes fabrication of a poly(dimethyl siloxane) (PDMS)-based chip to analyze multiple protein interactions utilizing glycidyl methacrylate (GMA) photopolymer for a site-specific immobilization of capture proteins in a closed system. First, using one direction channels of a PDMS mold having cross-channels, GMA micropads were prepared by photopolymerizing GMA solution by 365 nm light irradiation at predetermined positions. After the first mold was replaced with a second mold having higher height or directly without mold changing, capture proteins were allowed to be covalently immobilized onto the surface of the epoxide-activated GMA pads. Following immobilization, poly(ethylene glycol) diacrylate (PEG-DA) precursor was photopolymerized at specific regions to generate plugs for prevention of mixing between different sample injection channels, diminishing the need of a mold changing for sample injections. Final chip was assembled by connecting separated sample injection channels using a connector mold. The viability of this strategy was successfully demonstrated by simultaneous detection of two different antigen-antibody interactions.

Biocompatible Materials↗
PubMed2005

Oligonucleotide chip for the diagnosis of HNF-1 alpha mutations.

Mutations in HNF-1 alpha cause maturity-onset diabetes of the young (MODY) type 3, which is the most prevalent MODY subtype in most countries. In the present study, we investigated an oligonucleotide microchip for the detection of the known HNF-1 alpha mutations. We first optimized the coupling chemistries for covalent immobilization of allele-specific oligonucleotides on aldehyde (CHO)- and thiocyanate (NCS)-activated glass slides and compared their hybridization efficiencies. CHO-glass was found to provide a more favorable environment for hybridization than NCS-glass, whereas the binding capacity of NCS-glass for amine-activated oligonucleotide was much greater than with CHO-glass. We also investigated the effects of the length of the capture probes on the hybridized signals. To determine the presence of HNF-1 alpha mutations in a human sample, we prepared an oligonucleotide chip from selected mutation sites of exon2 from HNF-1 alpha. Cy3-labeled RNA target probes were obtained by in vitro transcription of promoter-tagged PCR products from a wild-type blood sample and subsequent fragmentation. Hybridization of the chip with the RNA target probes successfully identified all of the genotypes for the tested sites. This work demonstrates that oligonucleotide chip-based analysis is a good candidate for routine clinical testing for HNF-1 alpha mutations.

DNA Mutational Analysis↗
PubMed2005

Polydiacetylene (PDA)-based colorimetric detection of biotin-streptavidin interactions.

The natural binding of streptavidin (STA) for a small molecule, biotin, has made it a useful tool in specific targeting application, due to their most specific noncovalent biological interactions in nature. Here, we describe a polydiacetylene (PDA)-based colorimetric biosensor to detect the STA-biotin interactions. We synthesized two kinds of biotin-labeled 10,12-pentacosadiynoic acid (PCDA) monomers having 2(ethylene oxide) or 3(ethylene oxide) spacer and prepared biotin-modified PDA liposomes using them. Upon the addition of STA, the modified PDA liposomes showed a color change from blue to red in about 1h followed by their aggregation and precipitation. The aggregation was found to be due to the cross-linking between the PDA liposomes caused by the STA having four biotin binding sites, which was confirmed by their TEM analysis. Since the STA-biotin interactions are widely adopted for various biological detection systems; the biotin-modified PDA sensor developed in this study has enormous potentials for the development of various colorimetric biosensors.

Acetylene↗
PubMed2005

Array-based mutation detection of BRCA1 using direct probe/target hybridization.

We describe here an efficient microarray-based multiplex assay to detect Korean-specific mutations in breast cancer susceptibility gene BRCA1 using direct probe/target hybridization. Allele-specific oligonucleotides were covalently immobilized on an aldehyde-activated glass slide to prepare an oligonucleotide chip. From a wild-type sample, a two-step method was used to generate labeled multiplex polymerase chain reaction (PCR) amplification products of genomic regions containing the mutation sites. Amino allyl-dUTP, an amine-modified nucleotide, was incorporated during multiplex PCR amplifications and a monofunctional form of cyanine 3 dye was subsequently attached to the reactive amine group of the PCR products. Hybridization of the labeled PCR products to the oligonucleotide chip successfully identified all of the genotypes for the selected mutation sites. This work demonstrates that oligonucleotides chip-based analysis is a good candidate for efficient clinical testing for BRCA1 mutations when combined with the indirect strategy to prepare labeled target samples.

BRCA1 Protein↗
PubMed2005

Diagnosis of HNF-1alpha mutations on a PNA zip-code microarray by single base extension.

In the present study, we exploited the superior features of peptide nucleic acids (PNAs) to develop an efficient PNA zip-code microarray for the detection of hepatocyte nuclear factor-1alpha (HNF-1alpha) mutations that cause type 3 maturity onset diabetes of the young (MODY). A multi-epoxy linker compound was synthesized and used to achieve an efficient covalent linking of amine-modified PNA to an aminated glass surface. PCR was performed to amplify the genomic regions containing the mutation sites. The PCR products were then employed as templates in a subsequent multiplex single base extension reaction using chimeric primers with 3' complementarity to the specific mutation site and 5' complementarity to the respective PNA zip-code sequence on the microarray. The primers were extended by a single base at each corresponding mutation site in the presence of biotin-labeled ddNTPs, and the products were hybridized to the PNA microarray. Compared to the corresponding DNA, the PNA zip-code sequence showed a much higher duplex specificity for the complementary DNA sequence. The PNA zip-code microarray was finally stained with streptavidin-R-phycoerythrin to generate a fluorescent signal. Using this strategy, we were able to correctly diagnose several mutation sites in exon 2 of HNF-1alpha with a wild-type and mutant samples including a MODY3 patient. This work represents one of the few successful applications of PNA in DNA chip technology.

DNA Mutational Analysis↗
PubMed2005

Simple synthesis of hierarchically ordered mesocellular mesoporous silica materials hosting crosslinked enzyme aggregates.

Hierarchically ordered mesocellular mesoporous silica materials (HMMS) were synthesized using a single structure-directing agent. The mesocellular pores are synthesized without adding any pore expander; the pore walls are composed of SBA-15 type mesopores. Small-angle X-ray scattering revealed the presence of uniform pore structures with two different sizes. Using HMMS as a nanoscopic template, hierarchically ordered mesocellular mesoporous carbon (HMMC) and polymer (HMMP) materials were synthesized. HMMS was used as a host for enzyme immobilization. To improve the retention of enzymes in HMMS, we adsorbed enzymes, and then employed crosslinking using glutaraldehyde (GA). The resulting crosslinked enzyme aggregates (CLEAs) show an impressive stability with extremely high enzyme loadings. For example, 0.5 g alpha-chymotrypsin (CT) could be loaded in 1 g of silica with no activity decrease observed with rigorous shaking over one month. In contrast, adsorbed CT without GA treatment resulted in a lower loading, which further decreased due to continuous leaching of adsorbed CT under shaking. The activity of crosslinked CT aggregates in HMMS was approximately 10 times higher than that of the adsorbed CT, which represents a 74-fold increase in activity per unit weight of HMMS due to higher CT loading.

Adsorption↗
PubMed2005

Multifunctional drug delivery system using starch-alginate beads for controlled release.

Utilizing starch-containing alginate beads, a novel drug delivery system (DDS) was developed. With the starch inside, the composite bead could be dried in its original bead shape and handled in the dried state. By employing alginate multi-coating strategy on the starch-alginate beads, detained or controlled release was efficiently achieved and successfully demonstrated for a model peptide drug, L-phenylalanine. The initial latent time and release rate of the drug inside the beads were able to be controlled simply by varying the number of multi-coatings. While the latent time for the initial release was negligible for non-coated starch-alginate beads, the latent times of beads coated one, two, and four times increased to 15, 30, and 70 min, respectively. Furthermore, the alginate component of the composite beads could adsorb and remove heavy metals such as lead from the body. These multifunctional beads combined with the novel coating process will greatly benefit alginate gel-based DDS.

Alginates↗
PubMedDate not supplied

Sequential feeding of glucose and valerate in a fed-batch culture of Ralstonia eutropha for production of poly(hydroxybutyrate-co-hydroxyvalerate) with high 3-hydroxyvalerate fraction.

Several important properties of poly(3-hydroxybutyric-co-3-hydroxyvaleric acids) (P(3HB-co-3HV) depend mainly on the HV unit fraction of the copolymer. Sequential and simultaneous feeding of glucose and valerate were employed to produce P(3HB-co-3HV) in a fed-batch culture of Ralstonia eutropha, and the effects of feeding models on the cell growth, 3HV unit fraction, and copolymer productivity have been investigated. The sequential feeding of glucose and then valerate resulted in a cell density of 110.2 g/L, 3HV unit fraction of 62.7 mol %, and copolymer productivity of 0.56 g/(L.h), while the latter simultaneous feeding strategy never achieved the 3HV fraction of P(3HB-co-3HV) higher than 50%. A nuclear magnetic resonance study confirmed that the production of random copolymer P(3HB-co-3HV) with high 3HV unit fraction was possible even with sequential feeding of glucose and valerate.

Bioreactors↗