급성충소돌기염의 임상 : 병리학적 고찰

양기화,조준필,하영천,한진우,정학재,손수창,이성희,황재훈,신철호 최신의학사 1989 最新醫學 Vol.32 No.4

인슐린 비의존형 당뇨병 환자에서 혈장 Endothelin-1농도의 변화

신양수,조희충,김원식,국기용,김용화,정종훈,문철웅,배학연,양성훈 朝鮮大學校 附設 醫學硏究所 1992 The Medical Journal of Chosun University Vol.17 No.2

Endothelin(ET) is a 21-residue peptide originally isolated from the cultured porcine endothelial cells. There are at least three genes for endothelin:endothelin-1(ET-1), endothelin-2(ET-2), and endothelin-3(ET-3). Endothelins are present in various human biological fluids including plasma, urine, breast milk, and saliva and have been found elevated plasma ET concentrations in patients with diabetes mellitus, Patients undergoing maintenance hemodialysis due to chronic renal failure, patients with acute myocardial infarction, and patients with subarachnoid hemorrhage. Endothelial cell damage is suspected to occur in diabetic patients and may be one important cause of angiopathy, a major complication in diabetes mellitus. The elevation of ET in diabetic patients may be a marker of, and further exacerbate, their vascular disease. We measured the levels of ET-1 in plasma of 50 patients with non-insulin dependent diabetes mellitus(NIDDM) and 25 normal subjects by radioimmunoassay. The plasma ET-1 concentration (mean±S.D.) in NIDDM was 6.461 A2.510 f㏖/ ㎖, and was significantly higher than in normal subjects (4.567±1.155f㏖/㎖) (P<0.05). The plasma ET-1 concentration (mean±S.D.) in diabetic retinopathy group( 7.15±2.454 f㏖/ml) was significantly elevated than those in otherwise uncomplicated groups (5.348±2.390 f㏖/㎖)(P<0.01). The correlation between any clinical parameters and plasma ET-1 levels in NIDDM was not significant, In conclusion, this study suggest that the elevated levels of ET-1 in diabetic patients may be play a important role in the pathogenesis of diabetic complications.

Purification and gene Cloning of α-Amylase of Neturospora crassa

Yang, Chul Hak 한국균학회 1988 韓國菌學會誌 Vol.16 No.3

α-Amylase (EC. 3.2.1.1) of Neurospora crassa(ATCC 9279) was cloned in E. coli HB101 using shotgun method, and the enzymes isolated from both N. crassa and E. coli were compared. Chromosomal DNA isolated from the spores of N crassa was partially digested with Pst1 restriction endonuclease and rejoined to pBR322 which had been digested with the same enzyme. The resulting recombinant DNA were introduced into E. coli HB101 which had competency by treating with CaCl₂. As the result, about 8000 colonies which showed tetracycline resistance were selected and two of the colonies which had 13.5 Kb recombinant plasmid exhibit starch degrading activity on starch-containing plate when treated with D-cycloserine. α-Amylases from both N. crassa and E. coli were DEAE-cellulose ion exchange column chromatograhy and Bio-Gel P150 gel filtration colunmn. As the result, about 81.3 fold and 5.6 fold purification in specific activities were obtained respectively, and specific activities of the gel filtrates were 6.1 μ/㎎ and 85 μ/㎎ respectively. The properties of both enzymes were compared and they showed quite the similar pasterns in optimal temperature, optimal pH and had same molecular weight about 100,000 daltons on gel filtration method. Optimal temperatures for both enzymes were 70℃ and optimal pH were about 6 and 10.

Purification and Characterization of Farnesyl Protein Transferase from Bovine Testis

Yang, Chul Hak,Baik, Young Jin,Ryu, Kwon Yul 생화학분자생물학회 1977 BMB Reports Vol.28 No.3

Famesyl protein transferase involved in the first step of post-translational modification of p21^(ras) proteins transfers the famesyl moiety from famesyl pyrophosphate to a cysteine residue in p21^(ras) proteins. The enzyme was first purified 30,000-fold from bovine testis by use of 30∼50% ammonium sulfate fractionation, DEAE-Sephacel ion exchange chromatography, Sephacryl S-300 gel filtration chromatography, Sephacryl S-200 gel filtration chromatography, and hexapeptide (Lys-Lys-Cys-Val-Ile-Met) affinity chromatography. The molecular weight of the purified enzyme was estimated to be ∼100 kDa by gel filtration and SDS-polyacrylamide gels showed two closely spaced bands of ∼50 kDa protein. These indicate that the enryme consists of two nonidentical subunits, a and β, which have slightly different molecular weights. The enzyme was inhibited by hexapeptide (Lys-Lys-Cys-Val-Ile-Met), which acted as an alternative substrate that competed for famesylation. Kinetic analysis by measuring initial velocities showed that famesyl protein transferase is a very slow enzyme. EDTA-treated famesyl protein transferase showed little activity with Mg^(2+) or Zn^(2+) alone, but required both Mg^(2+) and Zn^(2+) for the catalytic activity.

Quantitative Assay for the Binding of Jun-Fos Dimer and Activator Protein-1 Site

Yang,Chul-Hak,Jun,Gyo,Hahm,Eun-Ryeong,Lee,Sangkyou,Park,Seyeon,Lee,Dug-Keun The Korea Science and Technology Center 1999 BMB Reports Vol.32 No.6

The Jun and Fos families of eukaryotic transcription factors form heterodimers capable of binding to their cognate DNA enhancer elements. We are interested in searching for inhibitors or antagonists of the binding of the Jun-Fos heterodimer to the activator protein-1 (AP-1) site. The basic-region leucine zipper (bZIP) domain of c-Fos was expressed as a fusion protein with glutathione S-transferase, and allowed to form a heterodimer with the bZIP domain of c-Jun. The heterodimer was bound to glutathione-agarose, to which were added radiolabeled AP-1 uncleotides. After thorough washing, the gel-bound radioactivity was counted. The assay is faster than the coventional electrophoretic mobility shift assay because the gel electrophoresis step and the autoradiography step are eliminated. Moreover, the assay is very sensitive, allowing the detection of picomolar quantities of nucleotides, and is not affected by up to 50% dimethylsulfoxide, a solvent for hydrophobic inhibitors. Curcumin and dihydroguaiaretic acid, recently known inhibitors of Jun-Fos-DNA complex formation, were applied to this Jun-GST-fused Fos system and revealed to decrease the dimer-DNA binding.

An Efficient System for the Expression and purification of Yeast Geranylgeranyl Protein Transferase Type Ⅰ

Yang, Chul-Hak,Kim, HyunKyung,Kim, Youngah The Korea Science and Technology Center 1998 BMB Reports Vol.31 No.1

To purify the geranylgeranyl protain transferase type Ⅰ(GGPT-I) efficiently, a gene expression system using the pGEX-4T-1 vector was constructed. The cal1 gene, encoding the βsubunit of GGPT-I, was subcloned into the pGEX-4T-1 vector and co-transformed into E. coli cells harboring the ram2 gene, the α subunit gene of GGPT-I. GGPT-I was highly expressed as a fusion protein with glutathione S-transferase (GST) in E. coli, purified to homogeneity by glutathione-agarose affinity chromatography, and the GST moiety was excised by thrombin treatment. The purified yeast GGPT-I showed a dose-dependent increase in the transferase activity, and its apparent K?? value for an undecapeptide fused with GST (GST-PEP) was 0.66 μM and the apparent K?? value for geranylgeranyl pyrophosphate (GGPP) was 0.071 μM.

Plenary Lecture and 2007 KSBMB Presidential Lecture : Negative regulation of β-catenin/Tcf signaling by natural products in gastric and colon cancer cell Lines

( Chul Hak Yang ) 한국생화학분자생물학회 (구 한국생화학회) 2007 생화학분자생물학회 춘계학술발표논문집 Vol.2007 No.-

Chemical Modification Studies of Yeast Farnesyl Protein Transferase

Yang, Chul Hak,Sohn, Seung Wan,Jun, Gyo 생화학분자생물학회 1998 BMB Reports Vol.30 No.4

Phenylglyoxal, diethyl pyrocarbonate (DEPC), and 1-cyclohexyl-3-[2-morpholinoethyl]-carbodiimide metho-p-toluenesulfonate (CMC) are modifying reagents specific for arginine, histidine, and aspartate or glutamate, respectively. They were found to inactivate S. cerevisiae farnesyl protein transferase (FPTase). The peptide substrate protected the enzyme against inactivation by CMC. and the other substrate farnesyl pyrophosphate showed protection against inactivation by phenylglyoxal. while neither of the two substrates protected the enzyme against DEPC inactivation. These results suggest the presence of aspartate/glutamate, arginine and histidine residues at the active site of this enzyme.

Quantitative Assay for the Binding of Jun-Fos Dimer and Activator Protein-1 Site

Yang, Chul Hak,Jun, Gyo,Lee, Dug Keun,Lee, Sang Kyou,Park, Se Yeon,Hahm, Eun Ryeong 생화학분자생물학회 1998 BMB Reports Vol.32 No.6

The Jun and Fos families of eukaryotic transcription factors form heterodimers capable of binding to their cognate DNA enhancer elements. We are interested in searching for inhibitors or antagonists of the binding of the Jun-Fos heterodimer to the activator protein-1 (AP-1) site. The basic-region leucine zipper (bZIP) domain of c-Fos was expressed as a fusion protein with glutathione S-transferase, and allowed to form a heterodimer with the bZIP domain of c-Jun. The heterodimer was bound to glutathione-agarose, to which were added radiolabeled AP-1 nucleotides. After thorough washing, the gel-bound radioactivity was counted. The assay is faster than the conventional electrophoretic mobility shift assay because the gel electrophoresis step and the autoradiography step are eliminated. Moreover, the assay is very sensitive, allowing the detection of picomolar quantities of nucleotides, and is not affected by up to 50% dimethylsulfoxide, a solvent for hydrophobic inhibitors. Curcumin and dihydroguaiaretic acid, recently known inhibitors of Jun-Fos-DNA complex formation, were applied to this Jun-GST-fused Fos system and revealed to decrease the dimer-DNA binding.

An Efficient System for the Expression and Purification of Yeast Geranylgeranyl Protein Transferase Type I

Yang, Chul Hak,Kim, Young Ah,Kim, Hyun Kyung 생화학분자생물학회 1999 BMB Reports Vol.31 No.1

To purify the geranylgeranyl protein transferase type I (GGPT-I) efficiently, a gene expression system using the pGEX-4T-1 vector was constructed. The cal 1 gene, encoding the β subunit of GGPT-I, was subcloned into the pGEX-4T-1 vector and co-transformed into E. coli cells harboring the ram2 gene, the α subunit gene of GGPT-I. GGPT-I was highly expressed as a fusion protein with glutathione S-transferase (GST) in E. coli, purified to homogeneity by glutathione-agarose affinity chromatography, and the GST moiety was excised by thrombin treatment. The purified yeast GGPT-I showed a dose-dependent increase in the transferase activity, and its apparent K_m value for an undecapeptide fused with GST (GST-PEP) was 0.66μM and the apparent K_m value for geranylgeranyl pyrophosphate (GGPP) was 0.071 μM.