• 제목/요약/키워드: SH3

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Molecular Cloning and NMR Characterization of the Nonreceptor Tyrosine Kinase PTK6 SH3-SH2-Linker Domain

  • Lee, Young-Min;Ahn, Kyo-Eun;Ko, Sung-Geon;Lee, Weon-Tae
    • Bulletin of the Korean Chemical Society
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    • 제30권5호
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    • pp.1043-1046
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    • 2009
  • Human protein tyrosine kinase-6 (PTK6) is a member of the non-receptor protein tyrosine kinase family and it is found in two-thirds of all breast tumors. Very recently, we proposed that the SH3 domain of PTK6 interacts with the linker region (Linker) between the SH2 and kinase domains, proving that the interaction between SH3 domain and Linker plays an important role in auto-inhibition mechanism. Residues from 1 to 191 corresponding region of SH3-SH2-Linker (SH32L) of PTK6 was cloned into the pET32a expression vector with Tobbaco etch virus (TEV) protease enzyme site by sequence homology and 3D structural model. The purified PTK6-SH32L was determined as a monomer conformation in solution. The amide proton resonances in the $^{15}N-^{1}H$ 2D-HSQC spectrum suggest that PTK6-SH32L possesses disordered structural region of the flexible/unstructured linker region. In addition, the backbone amide proton chemical shifts of the SH3 domain in the PTK6-SH32L differ from that of the independent domain, indicating that intra-molecular interaction between SH3 and Linker in the PTK6-SH32L is present.

Expression, Purification and NMR studies of SH3YL1 SH3 domain

  • Shrestha, Pravesh;Yun, Ji-Hye;Lee, Weon-Tae
    • 한국자기공명학회논문지
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    • 제14권2호
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    • pp.105-116
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    • 2010
  • SH3YL1, a novel protein containing one Src homology 3 domain at the carboxyl terminus was first detected in mouse anagen skin cDNA. This protein had a significant homology with YHRO 16c/Ysc 84, the yeast Src homology 3 domain-containing protein. The sequence identity was remarkable at the carboxyl and amino-terminal Src homology 3 domain, suggesting that the novel protein is a mouse homolog of the yeast protein and thus was termed as SH3YL1. SH3YL1 is composed of two domains, a DUF500 at N-termini and a SH3 domain at C-termini. In our study we cloned the SH3 domain in bacterial expression system in Escherichia coli using pET32a vector with TEV protease cleavage site and purified as a monomer using affinity chromatography. The N-terminal poly-Histidine tag was cleaved with TEV protease and target protein was used for backbone studies. Our study showed that SH3 domain primarily consists of $\beta$-sheet which is in consistence with previous result performed on the truncated SH3 domain of SH3YL1.

Functional analysis of SH3 domain containing ring finger 2 during the myogenic differentiation of quail myoblast cells

  • Kim, Si Won;Lee, Jeong Hyo;Park, Tae Sub
    • Asian-Australasian Journal of Animal Sciences
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    • 제30권8호
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    • pp.1183-1189
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    • 2017
  • Objective: Owing to the public availability of complete genome sequences, including avian species, massive bioinformatics analyses may be conducted for computational gene prediction and the identification of gene regulatory networks through various informatics tools. However, to evaluate the biofunctional activity of a predicted target gene, in vivo and in vitro functional genomic analyses should be a prerequisite. Methods: Due to a lack of quail genomic sequence information, we first identified the partial genomic structure and sequences of the quail SH3 domain containing ring finger 2 (SH3RF2) gene. Subsequently, SH3RF2 was knocked out using clustered regularly interspaced short palindromic repeat/Cas9 technology and single cell-derived SH3RF2 mutant sublines were established to study the biofunctional activity of SH3RF2 in quail myoblast (QM7) cells during muscle differentiation. Results: Through a T7 endonuclease I assay and genotyping analysis, we established an SH3RF2 knockout (KO) QM7#4 subline with 61 and 155 nucleotide deletion mutations in SH3RF2. After the induction of myotube differentiation, the expression profiles were analyzed and compared between regular QM7 and SH3RF2 KO QM7#4 cells by global RNA sequencing and bioinformatics analysis. Conclusion: We did not detect any statistically significant role of SH3RF2 during myotube differentiation in QM7 myoblast cells. However, additional experiments are necessary to examine the biofunctional activity of SH3RF2 in cell proliferation and muscle growth.

Purification and Spectroscopic Characterization of the Human Protein Tyrosine Kinase-6 SH3 Domain

  • Koo, Bon-Kyung;Kim, Min-Hyung;Lee, Seung-Taek;Lee, Weon-Tae
    • BMB Reports
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    • 제35권3호
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    • pp.343-347
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    • 2002
  • The human protein tyrosine kinase-6 (PTK6) polypeptide that is deduced from the cDNA sequence contains a Src homology (SH) 3 domain, SH2 domain, and catalytic domain of tyrosine kinase. We initiated biochemical and NMR characterization of PTK6 SH3 domain in order to correlate the structural role of the PTK6 using circular dichroism and heteronuclear NMR techniques. The circular dichroism data suggested that the secondary structural elements of the SH3 domain are mainly composed of $\beta$-sheet conformations. It is most stable when the pH is neutral based on the pH titration data. In addition, a number of cross peaks at the low-field area of the proton chemical shift of the NMR spectra indicated that the PTK6 SH3 domain retains a unique and folded conformation at the neutral pH condition. For other pH conditions, the SH3 domain became unstable and aggregated during NMR measurements, indicating that the structural stability is very sensitive to pH environments. Both the NMR and circular dichroism data indicate that the PTK6 SH3 domain experiences a conformational instability, even in an aqueous solution.

알팔파 재분화에서 배지조성에 따른 뿌리 유도율 조사 (Examination of Root Induction Ratio for Regeneration of Alfalfa by Medium Component)

  • 김기용;강경민;박근제;배은경;이인애;이병현;곽상수;조진기
    • 한국초지조사료학회지
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    • 제23권2호
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    • pp.95-100
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    • 2003
  • 알팔파 종자를 $3\;mg/{\ell}$의 2,4-D를 첨가한 SH 배지에서 배양하여 캘러스를 유도하였으며, 캘러스를 SH 배지에 $5\;mg/{\ell}$의 NAA와 $2\;mg/{\ell}$의 kinetin을 첨가한 배지에서 28~30일간 배양하고, $11\;mg/{\ell}$의 2,4-D와 $1\;mg/{\ell}$의 kinetin을 첨가한 배지에서 3~5일간 배양 후, $1.6\;g/{\ell}$의 (${NH_4}_2{SO_4}$$5.75\;g/{\ell}$의 proline을 첨가한 배지에서 21~25일간 배양하여 완전히 재분화된 식물체와 많은 수의 shoot를 얻었다. 알팔파의 재분화 과정에서 뿌리가 유도되는 효율을 높이기 위해, SH salt의 양과 호르몬 양을 달리한 8종의 배지에서 각각 뿌리가 유도되는 정도를 조사해 본 결과, SH 기본배지를 사용한 것에 비해 IBA를 첨가했을 때 뿌리 유도율이 높았으며, 배지 1 리터당 1.5 mg의 IBA를 첨가했을 때 부리 유도율이 56.0%로서 가장 높았다. 또한 SH salt의 양을 1/2로 줄이고 IBA 첨가량을 다르게 하여 뿌리 유도율을 조사했을 때, SH 기본배지를 사용한 것에 비해 SH salt의 양을 1/2로 줄이고 뿌리 유도율이 높아졌으며, IBA를 첨가했을 때 푸리 유도율이 높았다. 이상의 결과에서 알팔파의 뿌리를 유도하기 위한 배지는, IBA 첨가량을 $1.5\;mg/{\ell}$로 하고 SH salt의 양을 1/2로 줄여서 사용하는 것이 좋을 것으로 판단된다.

Propolis Inhibits Neurite Outgrowth in Differentiating SH-SY5Y Human Neuroblastoma Cells

  • Kim, Han Bit;Yoo, Byung Sun
    • Toxicological Research
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    • 제32권3호
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    • pp.239-243
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    • 2016
  • Propolis is a multicomponent, active, complex resinous substance collected by honeybees from a variety of plant sources. We have studied the effect of propolis on neurite outgrowth of SH-SY5Y human neuroblastoma cells induced to differentiate by all-trans-retinoic acid (RA). Propolis, at a concentration of $3{\mu}g/mL$, had no significant effect on the viability of differentiating SH-SY5Y cells. However, the neurite outgrowth of the differentiating SH-SY5Y cells treated with propolis ($0.3{\sim}3{\mu}g/mL$) for 48 hr was significantly inhibited in a dose-dependent manner. Treatment of RA-stimulated differentiating SH-SY5Y cells with 0.3 to $3{\mu}g/mL$ propolis resulted in decreased level of transglutaminase and 43-kDa growth-associated protein (GAP-43) in a dose-dependent manner. The results indicate that propolis is able to inhibit neurite outgrowth of differentiating SH-SY5Y cells.

감자 역병의 친환경 방제를 위한 키토산 제형의 살포 (Application of Chitosan Preparations for Eco-friendly Control of Potato Late Blight)

  • 장태현;김병섭
    • 식물병연구
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    • 제18권4호
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    • pp.338-348
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    • 2012
  • 감자역병은 감자재배에서 가장 중요한 병 중의 하나다. 감자역병의 친환경방제를 위하여 SH-1와 SH-2을 이용하여 감자역병균의 균사생장, 예방효과, 잎을 이용한 생물검정 및 포장시험을 통해 평가하였다. 실내에서 유묘를 이용한 시험에서 SH-1와 SH-2 제형을 살포 후 역병균을 접종 후 예방효과를 조사한 결과 95% 이상 발병을 억제하는 효과가 있었다. SH-1과 SH-2의 균사생장 억제 효과는 $300{\mu}g/ml$의 농도에서 균총의 크기를 74% 이상을 억제하였다. 잎을 이용한 생물검정에서 1% SH-1와 SH-2을 살포 후 1시간 뒤에 병원균을 접종한 결과 병의 발달을 줄였다. 감자 줄기에 총 폴리페놀 함량도 SH-1와 SH-2을 살포한 구에서 증가하였다. 포장에서 역병방제효과 시험에서 1% SH-1와 SH-2을 7일 간격 4회 살포한 처리구에서 각각 72%와 53%의 역병 방제 효과가 있었으며, 14일간 3회 살포한 구에도 유사한 병 방제효과를 보였다. 감자비대기에 1% SH-1와 SH-2 살포는 감자 생체 중을 증가시켰으며, 상품의 비율도 높았다. 이 결과에 의하면 SH-1와 SH-2의 살포는 친환경 유기농업에서 천연살균제로서 역병 방제에 사용 할 수 있으며, 감자 수확량도 증가시킬 수 있다.

Linear Alkylbenzene Sulfonate 분해세균의 분리 및 동정 (Isolation and Identification of Linear Alkylbenzene Sulfonate Degrading Bacteria)

  • 이기무;최우영
    • 농업과학연구
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    • 제21권1호
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    • pp.60-66
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    • 1994
  • 하수 및 토양시료로부터 다수의 LAS분해세균을 분리하고 고농도 (LAS 200ppm)의 액체 배지에서 생육하면서 분해력이 높은 KL-3, SH-2, EN-1등 3개 균주를 선발하였다. 이들의 형태적, 생리적 및 배양적 성질에 따라 KL-3은 Klebsiella속, SH-2는 Shigella속, EN-1은 Enterobacter속 균주로 각각 동정되었다. 실험실용 발효조에서의 LAS의 분해양상은 배양초기에 분해속도가 빠르고 (1 일후의 분해율: KL-3 및 SH-2, 50%; EN-1, 20%), 배양 1-2일사이에서 그 속도가 크게 지연되다가 다시 분해가 빠르게 진행되며 3, 4일후에는 다시 늦어지는 경향을 나타내었다 (7일후의 분해율: KL-3, 85%; SH-2, 82%; EN-1, 75%). 균체의 생육은 배양 2일 후에 $10^8cfu/mL$수준의 정지기에 도달하였으며 pH는 최초 7.0에서 6.2~6.7의 범위로 저하되어 서서히 변화하는 경향이었다. 균체에 의한 LAS의 흡착량은 Shigella sp. SH-2균주의 경우가 가장 많았으며 Freund1ich흡착등식 Y= 0.030X + 0.95를 나타내었다.

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한국응급구조학회지 게재 논문의 중심 단어 분석(2005년-2011년) (Coincidence analysis of keywords and MeSH terms in the Korean Journal of Emergency Medical Services)

  • 이경희;함영림
    • 한국응급구조학회지
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    • 제16권2호
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    • pp.43-51
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    • 2012
  • Purpose : We try to disclose how much the keywords of the papers from the Korean Journal of Emergency Medical Services with Medical Subject Headings(MeSH) terminologies and to understand the major subjects of the recent emergency medical technology research in Korea from keywords. Methods : We analyzed keywords from 524 articles of the Korean Journal of Emergency Medical Services that were published between 2005 and 2011. We investigated frequently used keywords and what percentages of keywords agree with MeSH terms using the MeSH browser. Results : There were on average 3.2 keywords per article. The most frequent key words were AED, Attitude, Cardiopulmonary Resuscitation, CPR, EMT, EMT students, External Defibrillator, Job satisfaction, Knowledge, 119 EMT in order. The number of terms in precise agreement with MeSH headings was 101(19.3%); 327 terms(62.4%) were not found in the MeSH browser and 96 terms(18.3%) partially matched MeSH terms. Conclusion : Many keywords used in the Korean Journal of Emergency Medical Services did not agree with MeSH terms. We conclude that contribution rules should be using MeSH terms and authors should be educated in the proper use of MeSH terms in their research and subsequent publication.

IBA 처리에 의한 버즈풋 트레포일 줄기 절단면에서 뿌리 분화 (Root Initiation in Cut Birdsfoot trefoil Stems by Treatment of IBA)

  • 김기용;최기준;성병렬;임용우;임영철;장요순;김원호
    • 한국초지조사료학회지
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    • 제21권1호
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    • pp.35-38
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    • 2001
  • 버즈풋 트레포일 (Lotus corniculatus L.)의 줄기부분을 절단하여, SH 배지에 IBA를 첨가한 농도별로 뿌리 유도 정도를 조사한 결과. IBA 농도가 높을수록 뿌리 유도가 좋았으며, 2.5 mg/$\ell$의 IBA를 첨가한 처리구에서 뿌리 유도 정도가 90~95%로서 가장 좋았다. 캘러스로부터 shoot를 유도한 다음 줄기를 취하여 처리한 경우에는, 처리 12일째에 1/2 SH-0 배지에서 9개 (45%), SH-0 배지에서 10 (50%), SH-0.5IBA 배지에서 10개 (50%), SH-1.0IBA 배지에서 10개 (50%), SH-1.5IBA 배지에서 13개 (65%), SH-2.0IBA 배지에서 14개 (70%), SH-2.5IBA 배지에서 19개 (95%). SH-3.0IBA 배지에서 14개 (70%)의 뿌리유도를 나타내었다.

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