• Journal of Microbiology and Biotechnology
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• v.17 no.8
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• pp.1308-1315
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• 2007

Lipase B from Candida antarctica (CalB) displayed on the cell surface of H. polymorpha has been functionally improved for catalytic activity by molecular evolution. CalB was displayed on the cell surface by fusing to a cell-wall anchor motif (CwpF). A library of CalB mutants was constructed by in vivo recombination in H. polymorpha. Several mutants with increased whole-cell CalB activity were acquired from screening seven thousand transformants. The two independent mutants CalB 10 and CalB 14 showed an approximately 5 times greater whole-cell activity than the wild-type. When these mutants were made as a soluble form, CalB 10 showed 6 times greater activity and CalB 14 showed an 11 times greater activity compared with the wild-type. Sequence analyses of mutant CALB genes revealed amino acid substitutions of $Leu^{278}Pro$ in CalB10 and $Leu^{278}Pro/Leu^{219}Gln$ in CalB14. The substituted $Pro^{278}$ in both mutants was located near the proline site of the ${\alpha}$10 helix. This mutation was assumed to induce a conformational change in the ${\alpha}$10 helix and increased the $k_{cat}$ value of mutant CalB approximately 6 times. Site-directed mutagenized CalB, LQ ($Leu^{219}Gln$) was secreted into the culture supernatant at an amount of approximately 3 times more without an increase in the CalB transcript level, compared with the wild-type.

• Bulletin of the Korean Chemical Society
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• v.26 no.12
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• pp.2033-2037
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• 2005

In vitro selection was used to isolate $Mg^{2+}$-dependent self-cleaving ribozymes with cis-cleavage activity from a pre-tRNA library having 40-mer random sequences attached to 5'-end of E. coli $tRNA^{Phe}$. After 8 rounds of SELEX (Systematic Evolution of Ligands by Exponential Enrichment), RNA molecules which can self-cleave at the high concentration of $Mg^{2+}$ were isolated. The selected ribozymes can carry out the self-cleavage reaction in the presence of 100 mM $Mg^{2+}$ but not in 10 mM $Mg^{2+}$. The cleavage sites of the ribozymes are located at +3 and +4 of $tRNA^{Phe}$, compared with +1 position of 5'-end cleavage site of pre-tRNA by RNase P. New RNA constructs deprived of its D stem-loop, anticodon stem-loop, variable loop and T stem-loop, respectively showed the cleavage specificity identical to a ribozyme having the intact tRNA structure. Also, the new ribozyme fused with both a ribozyme and $tRNA^{Leu}$ showed the cleavage activities at the various sites within its sequences, different from two sites of position +3 and +4 observed in the ribozyme with $tRNA^{Phe}$. Our results suggest that the selected ribozyme is not structural-specific for tRNA.

• Investigative Magnetic Resonance Imaging
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• v.12 no.2
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• pp.107-114
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• 2008

Purpose : The aim of this study is to determine possibility of application of in vivo proton ($^1H$) magnetic resonance spectroscopy (MRS) in distinguishing cystic mass arising around pancreas by comparison of in vivo MRS, in vitro MRS using 3T MR machine, based on nuclear magnetic resonance (NMR). Materials and Methods : We obtained spectra of in vivo MRS, in vitro MRS and NMR from abdominal mass arising around pancreas (mucinous cystic neoplasm=5, intraductal papillary mucin producing tumor=5, pseudocyst=1, and lymphangioma=1). We estimated existence of peak of in vivo MRS, and in vitro MRS concordant to that of NMR. We also evaluated differential peak for predicting specific disease. Results : Correlation of presence of peak with NMR showed showed sensitivity of 29.6%, specificity of 82.6% and accuracy of 67.7% on in vivo MRS (p = 0.096, McNemar test), sensitivity of 57.1% and specificity of 92.6% and accuracy of 82.3% on in vitro MRS (p = 0.362, McNemar test). The spectra of NMR for IPMT showed more frequent peaks at 3.5-4.0 ppm (p=0.026). Conclusion : Although chemical analysis, using NMR could be regarded as possible tool to differentiate cystic masses, in vivo and in vitro MRS need further technical evolution for clinical application.

• Development and Reproduction
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• v.15 no.3
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• pp.187-195
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• 2011

X chromosome inactivation (XCI) is a process that enables mammalian females to ensure the dosage compensation for X-linked genes. Investigating the mechanism of XCI might provide deeper understandings of chromosomal silencing, epigenetic regulation of gene expressions, and even the course of evolution. Studies on mammalian XCI conducted with mice have revealed many fundamental findings on XCI. However, difference of murine and human XCI necessitates the further investigation in human XCI. Recent success in reprogramming of differentiated cells into pluripotent stem cells showed the reversibility of XCI in vitro, X chromosome reactivation (XCR), which provides another tool to study the change in X chromosome status. This review summarizes the current knowledge of XCI during early embryonic development and describes recent achievements in studies of XCI in reprogramming process.

• The Korean Journal of Nuclear Medicine
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• v.38 no.2
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• pp.131-139
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• 2004

Small animal models are extensively utilized in the study of biomedical sciences. Current animal experiments and analysis are largely restricted to in vitro measurements and need to sacrifice animals to perform tissue or molecular analysis. This prevents researchers from observing in vivo the natural evolution of the process under study. Imaging techniques can provide repeatedly in vivo anatomic and molecular information noninvasively. Small animal imaging systems have been developed to assess biological process in experimental animals and increasingly employed in the field of molecular imaging studies. This review outlines the current developments in nuclear medicine imaging instrumentations including fused multi-modality imaging systems for small animal imaging.

• Biotechnology and Bioprocess Engineering:BBE
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• v.8 no.2
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• pp.64-75
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• 2003

Aptamers are functional nucleic acids that can specially bind to proteins, peptides, amino acids. nucleotides, drugs, vitamins and other organic and inorganic compounds. The aptamers are identified from random DNA or RNA libraries by a SELEX (systematic evolution of ligands by exponential amplification) process. As aptamers have the advantage, and potential ability to be released from the limitations of antibodies, they are attractive to a wide range of therapeutic and diagnostic applications. Aptamers, with a high-affinity and specificity, could fulfil molecular the recognition needs of various fields in biotechnology. In this work, we reviewed some aptamer Selection techniques, properties, medical applications of their molecules and their biotechnological applications, such as ELONA (enzyme linked oligonucleotide assay), flow cytometry, biosensors, electrophoresis, chromatography and microarrays.

• Journal of Microbiology and Biotechnology
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• v.16 no.7
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• pp.1149-1153
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• 2006

In this study, nuclease-resistant RNA aptamers that are specific for Jurkat T leukemia cells were selected by a subtractive systemic evolution of ligands by exponential enrichment (SELEX) method. A randomized nuclease-resistant RNA library was incubated with normal peripheral blood mononuclear cells (PBMC) in each round to preclude RNAs that recognize the common cellular components on the surface of normal and cancer cells. The precluded RNAs were used for the selection of Jurkat T cell-specific aptamers, and the specific RNAs were then gradually enriched from start to the following selections. After 16 rounds of the subtractive SELEX, the selected aptamers were found to preferentially bind to Jurkat T cells, but not to the normal PBMC, evidenced by fluorescence-activated cell sorting analysis. Thus, the subtractive SELEX can be used to identify ligands to cancer-specific biological markers without prior knowledge of the nature of markers. The aptamers could be applied to specific cell sorting, tumor therapy, and diagnosis, and moreover, to find cancer cell-specific markers.

• Genomics & Informatics
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• v.3 no.2
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• pp.77-80
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• 2005

Targeting of complex system such as human cells rather than biochemically pure molecules will be a useful approach to massively identify ligands specific for the markers associated with human disease such as cancer and simultaneously discover the specific molecular markers. In this study, we developed in vitro selection method to identify nuclease-resistant nucleic acid ligands called RNA aptamers that are specific for human cancer cells. This method is based on the combination of the cell-based selection and subtractive systematic evolution of ligands by exponential enrichment (SELEX) method. These aptamers will be useful for cancer-specific ligands for proteomic research to identify cancer-specific molecular markers as well as tumor diagnosis and therapy.

• Korean Journal of Materials Research
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• v.33 no.8
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• pp.337-343
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• 2023

3Y-TZP (3 mol% yttria-stabilized tetragonal zirconia polycrystals) ceramics have excellent mechanical properties including high fracture toughness, good abrasion resistance as well as chemical and biological stability. As a result, they are widely used in mechanical and medical components such as bearings, grinding balls, and hip implants. In addition, they provide excellent light transmittance, biocompatibility, and can match tooth color when used as a dental implant. Recently, given the materials' resemblance to human teeth, these ceramics have emerged as an alternative to titanium implants. Since the introduction of CAD/CAM in the manufacture of ceramic implants, they've been increasingly used for prosthetic restoration where aesthetics and strength are required. In this study, to improve the surface roughness of zirconia implants, we modified the 3Y-TZP surface with a biocomposite of hydroxyapatite and forsterite using room temperature spray coating methods, and investigated the mixed effect of the two powders on the evolution of surface microstructure, i.e., coating thickness and roughness, and biological interaction during the in vitro test in SBF solution. We compared improvement in bioactivity by observing dissolution and re-precipitation on the specimen surface. From the results of in vitro testing in SBF solution, we confirmed improvement in the bioactivity of the 3Y-TZP substrate after surface modification with a biocomposite of hydroxyapatite and forsterite. Surface dissolution of the coating layer and the precipitation of new hydroxyapatite particles was observed on the modified surface, indicating the improvement in bioactivity of the zirconia substrate.

• KSBB Journal
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• v.20 no.2 s.91
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• pp.88-92
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• 2005

Phosphorylation of amino acid residues in proteins, plays a major role in biological mechanism. Phosphorylation acts as a process regulating the protein activity in variable pathways such as metabolism, signal transduction and cell division. Therefore the development of ligands for phosphoamino acids are an important work for protein analysis and proteomics studies. In this study, RNA aptamers for o-phosphoserine, o-phosphotyrosine and o-phosphotyrosine which appears frequently in nature were developed by in vitro evolution method. We could obtain similar sequences from random RNAs of 40 mer by SELEX method through 10 cycles. As result, the aptamers for o-phosphoserine and o-phosphothreonine among phosphoamino acids aptamers showed high affinity of Kd=2.60 nM and 2.65 nM for their target molecules, respectively. In addition, these aptamers could be confirmed the high selectivity for their target.