• 한국동물학회지
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• 제37권1호
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• pp.40-48
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• 1994

세포분화에 따른 Small GTP-binding protein의 역할을 밝히기 위하여 Retinoic acid(RA)와 dibutyryl cyclic AMP(dbcAMP)로 분화를 유도한 F9 기형암종세포의 형태적인 변화와 함께 Small GTP-binding protein의 분포를 조사하였다. RA와 dbcAMP를 처리한 세포는 분화유도 5일경(초기 분화 단계)에 분명한 세포의 경계를 보이기 시작하여 7일경(분화 후기 단계)에는 거의 모든 세포가 등근 분화된 형태로 전환되었다. 이 분화과정 동안 세포막에는 많은 microvilli와 lamellopodia 같은 구조물이 나타났다. 아울러 초기 분화 단계에 많은 량의 laminin이 발현되었으며 분화 후기에 microtubule의 재분포가 관찰되었다. 세종류의 Small GTP-binding protein(25 23, 21 KD)이 F9 세포의 막성분과 세포질에서 관찰되었으며 분화가 진행됨에 따라서 세단백질 모두 증가되는 양상을 보였다 이러한 결과는 Small GTP-binding protein이 F9 세포의 분화에 특별한 기능을 가지고 있음을 시사해 주고 있다.

• BMB Reports
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• 제33권5호
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• pp.391-395
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• 2000

Calexcitin, a calcium-binding protein, was previously cloned and functionally characterized in the squid Loligo pealei. We now report the cloning of a second form of Calexcitin, Calexcitin-2, found in the squid Todarodes pacificus optic lobe. Calexcitin-2 has a significantly different carboxyl terminal region than Calexcitin-1. It lacks the CAAX motif, which is a farnesylation site. The amino acid sequence of Calexcitin-2 shows an 84% identity with Calexcitin-1 and also displays a strong cross immunoreactivity. Western blotting shows that Calexcitin-2 was expressed exclusively in the optic lobe region of squid, but not in other body organs. Regardless of its lack of conserved regions for GTP-binding, Calexcitin-2 shows moderately low affinity GTP-binding and also shows dramatic conformational change induced by GTP-binding. Three possible GTP-binding region mutations, K142A, D144A, and K157A, did not change the G TP binding affinity. This raises the possibility that Calexcitin-2 may have a novel GTP-binding motif.

• 한국동물학회지
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• 제39권3호
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• pp.292-298
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• 1996

안점의 꽃갯지렁이(Pseudopotamilla occelata)의 난자형성중 난황단백질의 전구체인 체강액 단백질(CP)은 수용체에 의해 중개되는데 이러한 수용체 중개에 의한 난모세포내로의 유입은 GTP-binding protein에 의해 조절되는 것으로 확인되었다. 체강액 단백질(CP)을 가장 활발히 투과시키는 중기 난모세포내로의 125 I-CP의 유입은 GTP에 의해 촉지되었고, GTP 유사체인 GTPrS나 GTP$\beta$S의 효과를 금입자로 표지된 체강액 단백질을 이용하여 전자현미경으로 확인해 본 결과, gold-labeled CP는 난황립에 집중되었고, 이러한 현상도 GTP에 의해서는 촉진되었고 GTPrS에 의해서는 억제되었다.

• Journal of Yeungnam Medical Science
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• 제10권2호
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• pp.360-368
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• 1993

세포막 정보전달 경로에서 수용체에 전달된 정보를 세포내로 전달하는데 조절 단백질로 알려진 GTP 결합단백질을 소외 뇌조직으로부터 정제하고 그 분자량등을 관찰하였다. 분쇄한 소의 뇌조직으로부터 세포막을 분리 해내고 1% cholate를 이용하여 세포막 단백질을 얻었으며 DEAE-Sephacel column chromatography를 시행하였다. 여기서 얻은 GTP 결합단백질은 다시 Ultrogel AcA 34 column chromatography, heptylamine-Sepharose column chromatography 순으로 실시하여 $GTP{\gamma}S$와 결합하는 단백질 분획을 모았고 활성도와 일치하는 분획을 얻었다. 전기영동으로 관찰한 결과 $Go{\alpha}$가 분자량 39,000 dalton. $G{\beta}$가 36,000 dalton인 band를 확인하였고 나머지 다른 단백질도 함께 관찰되어 heptylamine-Sepharose 분획을 다시 DEAE-Sephacel column에 적용하여 순수한 band를 구하였다. GTP 결합단백질의 활성화는 GTP가 결합될 때 ${\alpha}$부분과 결합하고 ${\beta}{\gamma}$는 떨어져 나간다. 그러므로 heptylamine-Sepharose column분획의 활성도에서 $Go{\alpha}$의 band 분획과 곡선의 활성도가 일치하고 ${\beta}$는 곡선이 하향하는 분획에서 전기영동상에 관찰되었다.

• 한국식물병리학회:학술대회논문집
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• 한국식물병리학회 2003년도 정기총회 및 추계학술발표회
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• pp.81.2-82
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• 2003

Plants have evolved along with pathogens, and they have developed sophisticated defense systems against specific microorganisms to survive. G-protons are considered one of the upstream signaling components working as a key for the defense signal transduction pathway. For activation and inactivation of G-protein, GTP-biding proteins are involved. GTP -binding proteins are found in all organisms. Small GTP-binding proteins, having masses of 21 to 30kD, belong to a superfamily, often named the Ras supefamily because the founding members are encoded by human Ras genes initially discovered as cellular homologs of the viral ras oncogene. Members of this supefamily share several common structural features, including several guanine nucleotide binding domains and an effector binding domain. However, exhibiting a remarkable diversity in both structure and function. They are important molecular switches that cycle between the GDP-bound inactive form into the GTP-bound active form through GDP/GTP replacement. In addition, most GTP-binding proteins cycle between membrane-bound and cytosolic forms. such as the RAC family are cytosolic signal transduction proteins that often are involved in processing of extracellular stimuli. Plant RAC proteins are implicated in regulation of plant cell architecture secondary wall formation, meristem signaling, and defense against pathogens. But their molecular mechanisms and functions are not well known. We isolated a RacB homolog from rice to study its role of defense against pathogens. We introduced the constitutively active and the dominant negative forms of the GTP-hinging protein OsRacB into the wild type rice. The dominant negative foms are using two forms (full-sequence and specific RNA interference with RacB). Employing southern, and protein analysis, we examine to different things between the wild type and the transformed plant. And analyzing biolistic bombardment of onion epidermal cell with GFP-RacB fusion protein revealed association with the nucle.

• Journal of Plant Biology
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• 제39권2호
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• pp.85-92
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• 1996

Small GTP-binding proteins are divided into three major group: Ras, Rho and Ypt/Rab. They have the conserved regions designed G1 to G5 that are critical in GDP/GTP exchange, GTP-induced conformational change and GTP hydrolysis. We isolated and characterized genomic DNA or cDNAfragments encoding G1 to G3 domains of small GTP-binding protein Rab and Rho from several plant species using two different PCR-based cloning strategies. Seven rab DNA fragments were isolated from 4 different plants, mung-bean, tobacco, rice and pepper using two degenerate primers corresponding to the GTP-binding domain G1 and G3 in small GTP-binding proteins. The amino acid sequences among these rab DNA fragments and other known small GTP-binding proteins shows that they belong to the Ypt/Rab family. Six rho DNA fragments were isolated from 5 different plants, mung-bean, rice, Arabidopsis, Allium and Gonyaulax using the nested PCR method that involves four degenerate primers corresponding to the GTP-binding domain G1, G3 and G4. The rho DNA fragments cloned show more than 90% homology to each other. Sequence comparison between plant and other known Rho family genes suggests that they are closely related (67 to 82% amino acid identity). Sequence analysis and southern blot analysis of rab and rho in mung-bean suggest than thses genes are encoded by multigene family in mung-bean.

• Journal of Photoscience
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• 제7권1호
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• pp.31-34
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• 2000

The heterotrichous ciliate Stentor coeruieus shows a step-up photophobic response to visible light In the previous paper, the existence of GTP-binding proteins was confirmed by using the antisera against the carboxy terminal decapeptide of transducin $\alpha$ subunit. The photoreceptor, stentorin, is localized in the pigment granule. If the immunoreactive G-protein directly interacts with the photoreceptor stentorin, the G-protein expected to be located in the pigment granule rather than plasma membrane. To elucidate the function of the immunoreactive G-protein, the localization of the G-protein in Stentor coeruleus was studied. The results suggest that this G-protein is located in the myoneme involved in the contraction and extension of the cell rather than in the pigment granule.

• Journal of Plant Biology
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• 제33권3호
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• pp.211-215
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• 1990

There is a family of homologous proteins known to small GTP-inding proteins which have a GTP binding domains and GTPase activity with molecular weight of about 20000 in mammalian tissues. Recently at least 20 different small GTP-binding proteins including three rasproto-oncogene, smg25, rho, and ral gene products were identified. These proteins play a central role in cellular prolifration, neoplasia, signal transduction, terminal differentiation, and secretory process of the cells. In this review, I have briefly compiled current information on the different areas of research in the small GTP-binding proteins in an attempt to convey an overall view of the fundamental role that this family of protein in normal cellular processes. Moreover, furture goals of research in the small GTP-binding proteins as well as the possible existence of this family of proteins in plant cells were discussed.

• BMB Reports
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• 제34권2호
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• pp.118-122
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• 2001

The translocation of ribose-binding protein (RBP) into the inverted membrane vesicles (IMV) of Escherichia coli and eukaryotic microsomes was studied using the in vitro translation/translocation system. It was found that RBP was translocated into heterologous eukaryotic microsomes co-translationally, as well as post-translationally However, RBP was translocated only past-translationally into IMV. Degradation fragments of RBP with the molar mass of 14 and 16 kDa were produced during the translocation into IMV However, the amount of the degradation products decreased and the mature form of RBP appeared in the presence of phenylmethylsulfonyl fluoride (PMSF). PMSF and GTP accelerated the translocation of RBF It was also found that SecB enhanced the post-translational translocation of RBP It appears that RBP is translocated via at least two targeting paths.

• 대한약리학회지
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• 제29권2호
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• pp.245-252
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• 1993

Amiloride는 $Na{^+}$ channels를 선택적으로 억제하는 potassium sparing diuretic이다. 본 연구에서는 amiloride와 아데노신 수용체의 상호작용을 밝히고자, 흰쥐에서 얻은 crude adipocytic membrane fractions의 adenylyl cyclase activity를 여러 조건하에서 측정하였다. 우선 GTP가 isoproterenol-stimulated adenylyl cyclase activity에 미치는 영향을 조사함으로서 $G_i$ protein (inhibitory guanine nucleotide binding protein)의 기능을 알아보았다. 그 결과 amiloride는 높은 GTP 농도에서 isoproterenol-stimulated adenylyl cyclase의 활성을 억제하는 것을 관찰할 수 없었다. 이와는 대조적으로 amiloride 존재 하에서 2-chloroadenosine을 사용하여 아데노신 수용체를 경유한 isoproterenol-stimulated adenylyl cyclase activity가 억제되는 정도를 측정하였을 때, 2-chloroadenosine의 농도에 따라 큰 변화 없거나 오히려 억제 효과가 더욱 크게 나타났다. 그러나 위와 같은 조건하에서 propranolol에 의한 isoproterenol-stimulated adenylyl cyclase activity의 억제는 amiloride에 의해서 유의하게 변하지 않는 것으로 보아서, 수용체를 매개로 한 $G_s$ protein의 기능은 amiloride에 의해 영향을 받지 않는 것으로 생각된다. 그리고 amiloride에 의해 증가된, 2-chloroadenosine-mediated adenylyl cyclase의 억제 효과는 150mM NaCl 존재 하에서도 그대로 유지되었다. 이러한 결과로 보아 amiloride는 아데노신 수용체와 결합하여 $G_i$ proteins과의 coupling을 용이하게 할 뿐만 아니라, $G_i$ protein을 선택적으로 변화시켜 $G_i$ protein의 GTP 의존적인 adenylyl cyclase의 억제 기능을 제거하는 두 작용을 갖는 것으로 사료된다.