• Title/Summary/Keyword: domain inclusion

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ON THE SHAPE DERIVATIVE IN THE DOMAIN INCLUSION

  • Kim, Hongchul
    • Korean Journal of Mathematics
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    • v.10 no.1
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    • pp.75-87
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    • 2002
  • The shape derivative for the domain functional will be discussed in the situation of domain inclusion. Hadamard's shape structure is sought by using the material derivative in conjunction with the domain imbedding technique.

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A micromechanics-based time-domain viscoelastic constitutive model for particulate composites: Theory and experimental validation

  • You, Hangil;Lim, Hyoung Jun;Yun, Gun Jin
    • Advances in aircraft and spacecraft science
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    • v.9 no.3
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    • pp.217-242
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    • 2022
  • This paper proposes a novel time-domain homogenization model combining the viscoelastic constitutive law with Eshelby's inclusion theory-based micromechanics model to predict the mechanical behavior of the particle reinforced composite material. The proposed model is intuitive and straightforward capable of predicting composites' viscoelastic behavior in the time domain. The isotropization technique for non-uniform stress-strain fields and incremental Mori-Tanaka schemes for high volume fraction are adopted in this study. Effects of the imperfectly bonded interphase layer on the viscoelastic behavior on the dynamic mechanical behavior are also investigated. The proposed model is verified by the direct numerical simulation and DMA (dynamic mechanical analysis) experimental results. The proposed model is useful for multiscale analysis of viscoelastic composite materials, and it can also be extended to predict the nonlinear viscoelastic response of composite materials.

Effects of Co-Expression of Liver X Receptor β-Ligand Binding Domain with its Partner, Retinoid X Receptor α-Ligand Binding Domain, on their Solubility and Biological Activity in Escherichia coli

  • Kang, Hyun
    • Journal of Microbiology and Biotechnology
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    • v.25 no.2
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    • pp.247-254
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    • 2015
  • In this presentation, I describe the expression and purification of the recombinant liver X receptor β-ligand binding domain proteins in E. coli using a commercially available double cistronic vector, pACYCDuet-1, to express the receptor heterodimer in a single cell as the soluble form. I describe here the expression and characterization of a biologically active heterodimer composed of the liver X receptor β-ligand binding domain and retinoid X receptor α-ligand binding domain. Although many of these proteins were previously seen to be produced in E. coli as insoluble aggregates or "inclusion bodies", I show here that as a form of heterodimer they can be made in soluble forms that are biologically active. This suggests that co-expression of the liver X receptor β-ligand binding domain with its binding partner improves the solubility of the complex and probably assists in their correct folding, thereby functioning as a type of molecular chaperone.

The Development of Social Inclusion.Exclusion Indicators - Focused on the Permanent Rental Apartment Dwellers - (사회적 통합.배제 지표 개발을 위한 연구 - 영구임대아파트 거주자를 중심으로 -)

  • Kim, Mi-Hee;Lee, Min-A;Noh, Se-Hee
    • Journal of the Korean housing association
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    • v.19 no.6
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    • pp.95-104
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    • 2008
  • This study was to develop the indicators for understanding social inclusion exclusion of the dwellers living in permanent rental apartment, and to present a important base about priority order of national housing policy for social inclusion. The ultimate purpose of this study was to provide basic information for the development of permanent rental apartment renewal techniques. The first phase of the study was to review of the social inclusion exclusion indicators mentioned in the literature. The indicators of EU (2001, 2006), KIHASA (2005), and Jehoel-Gijsbers & Brooman (2007), which were applied in many studies about social inclusion, or included various items about dwellers' subjective attitudes, were selected to construct the framework for the study. On the basis of 3 kinds of indicators at the above, the dimensions of social inclusion exclusion were categorized as material deprivation and access to social rights in an economicstructural exclusion view, and social participation and cultural normative accommodations in a socio-cultural exclusion view. And then, the domains of social inclusion exclusion were decided as follows: income, employment, education service, housing, health, family networks and social networks. The detail contents of indicators were adopted from the prior studies as many as possible, and the dwellers' subjective attitudes and housing domains were intensified with reference to UN housing rights and the study of "residents' satisfaction of housing facilities living in permanent rental housing". The developed indicators were modified through the advisory committee that consist of the specialists from the various fields of studies. The final indicators that were overlapped or not able to be measured were eliminated, and added, in a housing domain, the standards of convenient facilities, the management condition, safety, location, crime and etc. in the apartment complex, which were required to complement in the advisory committee.

Domain Expression of ErmSF, MLS (macrolide-lincosamide-streptogramin B) Antibiotic Resistance Factor Protein (MLS (macrolide-lincosamide-streptogramin B) 항생제 내성인자 단백질인 ErmSF의 domain발현)

  • 진형종
    • Korean Journal of Microbiology
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    • v.37 no.4
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    • pp.245-252
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    • 2001
  • Erm proteins, MLS (macrolide-lincosamide-streptogramin B) resistance factor proteins, show high degree of amino acid sequence homology and comprise of a group of structurally homologous N-methyltransferases. On the basis of the recently determined structures of ErmC` and ErmAM, ErmSF was divided into two domains, N-terminal end catalytic domain and C-terminal end substrate binding domain and attempted to overexpress catalytic domain in E. coli using various pET expression systems. Three DNA fragments were used to express the catalytic domain: DNA fragment 1 encoding Met 1 through Glu 186, DNA fragment 2 encoding Arg 60 to Glu 186 and DNA fragment 3 encoding Arg 60 through Arg 240. Among the pET expression vectors used, pET 19b successfully expressed the DNA fragment 3 and pET23b succeeded in expression of DNA fragment 1 and 2. But the overexpressed catalytic domains existed as inclusion body, a insoluble aggregate. To assist the soluble expression of ErmSF catalytic domains, Coexpression of chaperone GroESL or Thioredoxin and lowering the incubation temperature to $22^{\circ}C$ were attempted, as did in the soluble expression of the whole ErmSF protein. Both strategies did not seem to be helpful. Solubilization with guanidine-HCl and renaturation with gradual removal of denaturant and partial digestion of overexpressed whole ErmSF protein (expressed to the level of 126 mg/ι culture as a soluble protein) with proteinase K, nonspecific proteinase are under way.

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RRM but not the Asp/Glu domain of hnRNP C1/C2 is required for splicing regulation of Ron exon 11 pre-mRNA

  • Moon, Heegyum;Jang, Ha Na;Liu, Yongchao;Choi, Namjeong;Oh, Jagyeong;Ha, Jiyeon;Kim, Hyeon Ho;Zheng, Xuexiu;Shen, Haihong
    • BMB Reports
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    • v.52 no.11
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    • pp.641-646
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    • 2019
  • The Ron proto-oncogene is a human receptor for macrophage-stimulating protein (MSP). The exclusion of exon 11 in alternative splicing generates ${\Delta}RON$ protein that is constitutively activated. Heterogenous ribonucleaoprotein (hnRNP) $C_1/C_2$ is one of the most abundant proteins in cells. In this manuscript, we showed that both hnRNP $C_1$ and $C_2$ promoted exon 11 inclusion of Ron pre-mRNA and that hnRNP $C_1$ and hnRNP $C_2$ functioned independently but not cooperatively. Moreover, hnRNP $C_1$ stimulated exon 11 splicing through intron 10 activation but not through intron 11 splicing. Furthermore, we showed that, whereas the RRM domain was required for hnRNP $C_1$ function, the Asp/Glu domain was not. In conclusion, hnRNP $C_1/C_2$ promoted exon 11 splicing independently by stimulating intron 10 splicing through RRM but not through the Asp/Glu domain.

Soluble Expression and Purification of Human Tissue-type Plasminogen Activator Protease Domain

  • Lee, Hak-Joo;Im, Ha-Na
    • Bulletin of the Korean Chemical Society
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    • v.31 no.9
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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.

Solid-phase Refolding of Poly-lysine Tagged Fusion Protein of hEGF and Angiogenin

  • Park Sang Joong;Ryu Kang;Suh Chang Woo;Chai Young Gyu;Kwon Oh Byung;Park Seung Kook;Lee Eun Kyu
    • Biotechnology and Bioprocess Engineering:BBE
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    • v.7 no.1
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    • pp.1-5
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    • 2002
  • A fusion protein, consisting of a human epidermal growth factor (hEGF) as the recognition domain and human angiogenin as the toxin domain, can be used as a targeted therapeutic against breast cancer cells among others. The fusion protein was expressed as inclusion body in recombinant E. coli, and when the conventional, solution-phase refolding process was used the refolding yield was very low due to severe aggregation. It was probably because of the opposite electric charge at a neutral pH resulting from the vastly different pI values of each domain. The solid-phase refolding process that exploited the ionic interactions between ionic exchanger surface and the fusion protein was tried, but the adsorption yield was also very low, below $ 30\%$, regardless of the resins and pH conditions used. Therefore, to provide a higher ionic affinity toward the solid matrix, six lysine residues were tagged to the N-terminus of the hEGF domain. When heparin-Sepharose was used as the matrix, the adsorption capacity increased 2.5-3 times to about $88\%$. Besides the intrinsic affinity of angiogenin to heparin, the poly-lysine tag provided additional ionic affinity. And the subsequent refolding yield increased nearly 13-fold, from ca. $4.8\%$ in the conventional refolding of the untagged fusion protein to $63.6\%$. The process was highly reproducible. The refolded protein in the column eluate retained RNase bioactivity of angiogenin.

A Divalent Immunotoxin Formed by the Disulfide Bond between Hinge Regions of Fab Domain

  • Choe, Seong Hyeok;Kim, Ji Eun;Lee, Yong Chan;Jang, Yeong Ju;Choe, Mu Hyeon
    • Bulletin of the Korean Chemical Society
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    • v.22 no.12
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    • pp.1361-1365
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    • 2001
  • Recombinant immunotoxins are hybrid cytotoxic proteins designed to selectively kill cancer cells. A divalent immunotoxins, [B3(FabH1)-PE38]2, was constructed by recombining Fab domain of B3 antibody as a cell-targeting domain and Pseudo monas exotoxin A (PE) as a cytotoxic domain. Monoclonal antibody, B3, is the murine antibody (IgG1k) directed against Lewis Y-related carbohydrate antigens, which are abundant on the surface of many carcinomas. Fab fragment of this antibody was used in this study with the modified hinge sequence where last two cysteines out of three were mutated to serine. PE is a 66 kDa bacterial toxin that kills eukaryotic cells by inhibiting protein synthesis with ADP ribosylation of ribosomal elongation factor 2 (EF2). Fc region of B3 antibody was substituted with the truncated form of PE (38 kDa, PE38) on DNA level. [B3(FabH1)-PE38]2 was formed by disulfide bond between cysteines in the modified hinge region of B3(FabH1)-PE38. Each polypeptide for recombinant immunotoxins was overexpressed in Escherichia coli and collected as inclusion bodies. Each inclusion body was solubilized and refolded, and cytotoxic effects were measured. Divalent immunotoxins, [B3(FabH1)-PE38]2, had ID50 values of about 10 ng/mL on A431 cell lines and about 4 ng/mL on CRL1739 cell lines. Control immunotoxins, B3(scFv)-PE40, had ID50 values of about 28 ng/mL on A431 cell lines and about 41 ng/mL on CRL1739 cell lines. Divalent immunotoxins, [B3(FabH1)-PE38]2, had higher cytotoxic effects than B3(scFv)-PE40 control immunotoxins.