• 생명과학회지
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• 제16권3호
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• pp.415-419
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• 2006

방향족 에폭사이드 기질에 대한 입체선택적 가수분해능이 우수한 Rhodotorula glutinis의 epoxide hydrolase (EH)를 codon usage를 고려한 Escherichia coli 균주에서 고효율로 발현할 수 있었다. 효모인 R. glutinis와 박테리아인 E. coli에서의 codon usage 선호도를 분석하고 그 차이를 고려하여 E. coli 에서 잘 사용되지 않는 rare codon에 대한 tRNA유전자정보가 들어 있는 pRARE plasmid를 함유한 E. coli 균주인 Rosetta(DE3)pLysS를 숙주세포로 사용하였다. R. glutinis EH를 발현시킨 재조합 E. coli를 생촉매로 사용하여 라세믹 styrene oxide 혼합물과 반응시켰을 때, (R)-styrene oxide에 대한 입체선택적 가수분해활성이 wild type R. glutinis 대비 매우 향상됨을 관찰할 수 있었다. 또한 라세믹 기질로부터 입체적으로 고순도인 99% ee 값을 갖는 광학적으로 순수한 (S)-styrene oxide를 얻을 수 있었다.

• 공업화학
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• 제17권5호
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• pp.557-560
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• 2006

Aspergillus niger LK의 epoxide hydrolase (EH)를 codon usage를 고려한 Escherichia coli 균주에서 고효율로 발현할 수 있었다. E. coli에서 잘 사용되지 않는 rare codon에 대한 tRNA 유전자 정보가 들어있는 plasmid를 함유한 E. coli 균주인 Rosetta (DE3)PLysS를 숙주세포로 사용하였다. A. niger EH를 발현시킨 재조합 E. coli를 생촉매로 사용하여 라세믹 styrene oxide 혼합물과 반응시켰을 때, (R)-styrene oxide에 대한 입체선택적 가수분해활성이 향상됨을 확인할 수 있었다. 또한 라세믹 기질로부터 입체적으로 고순도인 99% ee 값을 갖는 광학적으로 순수한 (S)-styrene oxide를 얻을 수 있었다.

• The Plant Pathology Journal
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• 제21권4호
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• pp.334-342
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• 2005

The prototype Chlorella virus PBCV-l encodes 11 tRNA genes and over 350 protein-encoding genes in its 330 kbp genome. Initial attempts to overexpress the recombinant A189/192R protein, a putative virus attachment protein, in E. coli strain BL21(DE3) SI were unsuccessful, and multiple protein bands were detected on Western blots. However, the full-length A189/192R recombinant protein or fragments derived from it were detected when they were expressed in E. coli BL21 CodonPlus (DE3) RIL, which contains extra tRNAs. Codon usage analysis of the a189/192r gene showed highly biased usage of the AGA and AVA codons compared to genes encoded by E. coli and Chlorella. In addition, there were biases of XXA/U($56\%$) and XXG/ C($44\%$) in the codons recognized by the viral tRNAs, which correspond to the codon usage bias in the PBCV-1 genome of XXA/U ($63\%$) over those ending in XXC/G ($37\%$). Analysis of the codon usage in the major capsid protein and DNA polymerase showed preferential usage of codons that can be recognized by the viral tRNAs. The Asn (AAC) and Lys (AAG) codons whose corresponding tRNA genes are duplicated in the tRNA gene cluster were the most abundant (i.e., preferred) codons in these two proteins. The tRNA genes encoded in the PBCV-l genome seem to play a very important role during the synthesis of viral proteins through supplementing the tRNAs that are frequently used in viral proteins, but are rare in the host cells. In addition, these tRNAs would help the virus to adapt to a wide range of hosts by providing tRNAs that are rare in the host cells.

• Parasites, Hosts and Diseases
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• 제54권6호
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• pp.803-807
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• 2016

Toxascaris leonina is a common parasitic nematode of wild mammals and has significant impacts on the protection of rare wild animals. To analyze population genetic characteristics of T. leonina from South China tiger, its mitochondrial (mt) genome was sequenced. Its complete circular mt genome was 14,277 bp in length, including 12 proteincoding genes, 22 tRNA genes, 2 rRNA genes, and 2 non-coding regions. The nucleotide composition was biased toward A and T. The most common start codon and stop codon were TTG and TAG, and 4 genes ended with an incomplete stop codon. There were 13 intergenic regions ranging 1 to 10 bp in size. Phylogenetically, T. leonina from a South China tiger was close to canine T. leonina. This study reports for the first time a complete mt genome sequence of T. leonina from the South China tiger, and provides a scientific basis for studying the genetic diversity of nematodes between different hosts.

• Asian Pacific Journal of Cancer Prevention
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• 제14권7호
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• pp.4279-4282
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• 2013

Background: The activation and inactivation of receptor tyrosine kinases are tightly regulated to ensure faithful replication of cells. After having transduced extracellular growth activating signals, activated EGFR is subjected to downregulation either by clathrin mediated endocytosis or c-Cbl mediated proteasome degradation depending on the ligand concentration. c-Cbl is an ubiquitin ligase which requires a phosphorylated tyrosine residue at position 1045 in the cytoplasmic domain of EGFR to interact and add ubiquitin molecules. While activating mutations in exons 19 and 21 have been associated with the development of several cancers, the status of mutations at tyrosine 1045 coding exon 27 of EGFR remain to be investigated. Consistently, defective phosphorylation at 1045 has been associated with sustained phosphorylation of EGFR in non-small lung carcinomas. Hence in the present study we investigated the genetic status of the tyrosine 1045 coding site within exon 27 of EGFR gene to explore for possible occurrence of mutations in this region, especially since no studies have addressed this issue so far. Materials and Methods: Tumor chromosomal DNA isolated from thirty five surgically excised oral squamous cell carcinoma tissues was subjected to PCR amplification with intronic primers flanking the tyrosine 1045 coding exon 27 of EGFR gene. The PCR amplicons were subsequently subjected to direct sequencing to elucidate the mutation status. Results: Sequence analysis identified no mutations in the tyrosine 1045 codon of EGFR in any of the thirty five samples that were analyzed. Conclusions: The lack of identification of mutation in the tyrosine 1045 codon of EGFR suggests that mutations in this region may be relatively rare in oral squamous cell carcinomas. To the best of our knowledge, this study is the first to have explored the genetic status of exon 27 of EGFR in oral squamous cell carcinoma tissue samples.

• Bulletin of the Korean Chemical Society
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• 제31권9호
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• pp.2607-2612
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• 2010

Human tissue-type plasminogen activator (tPA) is a valuable thrombolytic agent used to successfully treat acute myocardial infarction, thromboembolic stroke, peripheral arterial occlusion, and venous thromboembolism. Recombinant tPA is accumulated as an inactive form in inclusion bodies of E. coli and is refolded in vitro, which is accompanied by extensive aggregation. In the present study, a tPA protease domain was expressed in an active soluble form in the cytosol of E. coli Rosetta-gami cells, which allowed disulfide bond formation and supplied the tRNA molecules required for six rarely used codons in E. coli. This strategy increased the amount of soluble protease domain protein and avoided the cumbersome refolding process. The purified protease domain not only degraded tPA substrate peptides but also formed a covalently bound complex with plasminogen activator inhibitor-1, as does full-length tPA. Soluble expression and purification of tPA domains may aid in functional analyses of this multi-domain protein, which has been implicated in many physiological and pathological processes.

• Clinical and Experimental Pediatrics
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• 제50권10호
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• pp.1024-1029
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• 2007

Costello 증후군은 특징적 얼굴 형태(coarse face), 발달 장애, 지능 저하, 성장 지연, 신경학적 이상, 심근증, 피부 병변, 수유 장애, 상대적 대두증, 소관절의 과신전, 고형종 발생 증가 등을 특징으로 하는 드문 증후군으로 1977년 Costello에 의해 처음으로 보고되었으며, 이후 전세계적으로 약 100례 이상이 보고되었다. 최근 Costello 증후군은 HRAS 유전자 내의 glycine 12 또는 13 codon을 침범하는 de novo mutation에 의해 발생하는 것으로 알려져 있다. 저자들은 특징적인 얼굴 형태와 지능 저하, 대두증, 손과 발의 과도한 주름, 비후성 심근증과 심방성 빈맥으로 특징지어지는 Costello 증후군으로 진단된 환아 3례를 경험하였기에 이를 보고하는 바이다.

• Journal of Microbiology and Biotechnology
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• 제26권2호
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• pp.356-363
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• 2016

Human papillomavirus (HPV), a non-enveloped, double-stranded DNA tumor virus, is a primary etiological agent of cervical cancer development. As a potential tool for prophylactic vaccination, the development of virus-like particles (VLPs) containing the HPV16 L1 capsid protein is highly desired. In this study, we developed a high-level expression system of the HPV16 L1 in Escherichia coli for the purpose of VLP development. The native gene of HPV16 L1 has many rare codons that cause the early termination of translation and result in the production of truncated forms. First, we optimized the codon of the HPV16 L1 gene to the preferable codons of E. coli, and we succeeded in producing the full-size HPV16 L1 protein without early termination. Next, to find the best host for the production of HPV16 L1, we examined a total of eight E. coli strains, and E. coli BL21(DE3) with the highest yield among the strains was selected. With the selected host-vector system, we did a fed-batch cultivation in a lab-scale bioreactor. Two different feeding solutions (complex and defined feeding solutions) were examined and, when the complex feeding solution was used, a 6-fold higher production yield (4.6 g/l) was obtained compared with that with the defined feeding solution.

• International Journal of Industrial Entomology and Biomaterials
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• 제46권2호
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• pp.41-54
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• 2023

The blue-tailed damselfly, Ischnura elegans Van der Linden, 1820 (Odonata: Coenagrionidae), is a climate-sensitive indicator species in South Korea. In this study, we sequenced the complete mitochondrial genome (mitogenome) of I. elegans collected from South Korea for subsequent population genetic analysis, particularly to trace population movements in response to climate change. The 15,963 base pair (bp)-long complete mitogenome of I. elegans has typical sets of genes including a major non-coding region (the A+T-rich region), and an arrangement identical to that observed in ancestral insect species. The ATP6, ND3 and ND1 genes have the TTG start codon, which, although rare, is the canonical start codon for animal mitochondrial tRNA. The A/T content was 71.4% in protein-coding genes, 72.1% in tRNAs, 72.9% in the whole genome, 74.7% in srRNA, 75.3% in lrRNA, and 83.8% in the A+T-rich region. The A+T-rich region is unusually long (1,196 bp) and contains two subunits (192 bp and 176-165 bp), each of which is tandemly triplicated and surrounded by non-repeat sequences. Comparison of the sequence divergence among available mitogenomes of I. elegans, including the one from the current study, revealed ND2 as the most variable gene, followed by COII and COI, suggesting that ND2 should be targeted first in subsequent population-level studies. Phylogenetic reconstruction based on all available mitogenome sequences of Coenagrionidae showed a strong sister relationship between I. elegans and I. senegalensis.

• Asian Pacific Journal of Cancer Prevention
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• 제17권2호
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• pp.603-608
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• 2016

Background: The investigation of mutation patterns in oncogenes potentially can make available a reliable mechanism for management and treatment decisions for patients with colorectal cancer (CRC). This study concerns the rate of KRAS and BRAF genes mutations in Iranian metastatic colorectal cancer (mCRC) patients, as well as associations of genotypes with clinicopathological features. Materials and Methods: A total of 1,000 mCRC specimens collected from 2008 to 2012 that referred to the Mehr Hospital and Partolab center, Tehran, Iran enrolled in this cross sectional study. Using HRM, Dxs Therascreen and Pyrosequencing methods, we analyzed the mutational status of KRAS and BRAF genes in these. Results: KRAS mutations were present in 33.6% cases (n=336). Of KRAS mutation positive cases, 85.1% were in codon 12 and 14.9% were in codon 13. The most frequent mutation at KRAS codon 12 was Gly12Asp; BRAF mutations were not found in any mCRC patients (n=242). In addition, we observed a strong correlation of KRAS mutations with some clinicopathological characteristics. Conclusions: KRAS mutations are frequent in mCRCs while presence of BRAF mutations in these patients is rare. Moreover, associations of KRAS genotypes with non-mucinous adenocarcinoma and depth of invasion (pT3) were remarkable.