• 한국식물병리학회지
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• 제2권1호
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• pp.37-42
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• 1986

[ $0.5\%$ ] lactophenol acid fuchsin으로 염색해 본 결과 보리흰가루 병균의 제1차 발아관에 반응하여 보리표피 세포에 papillae 및 cytoplasmic aggregate가 형성되었으나 그 크기는 부착기에 반응하여 형성된 것보다는 훨씬 작았으며 또한 접종후 36-48시간 이후의 표피세포내에는 acid fuchsin에 의해서 다른 세포들에 비해 세포전체가 좀 더 진하게 염색되는 곳이 군데군데 관찰되었다. 그러나 접종 후 96시간까지 지질과산화산물의 일종인 malondialdehyde의 함량은 증가하지 않았다. 접종 후 6시간에 형성된 papillae 및 cytoplasmic aggregate내에 callose, protein, phenol물질 등은 집적되었으나 접종 후 72시간까지도 cellulose, cutin, suberin, lignin등은 검출되지 않았다. 보리-흰가루병 상호조합의 상기 모든 반응은 race 비특이적이었다.

• Bulletin of the Korean Chemical Society
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• 제35권9호
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• pp.2781-2786
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• 2014

Human ${\alpha}_1$-antitrypsin (${\alpha}_1$-AT) is a natural inhibitor of neutrophil elastases and has several dozens of genetic variants. Most of the deficient genetic variants of human ${\alpha}_1$-AT are unstable and cause pulmonary emphysema. However, the most clinically significant variant, Z-type ${\alpha}_1$-AT, exhibits retarded protein folding that leads to accumulation of folding intermediates. These aggregate within the endoplasmic reticulum (ER) of hepatocytes, subsequently causing liver cirrhosis as well as emphysema. Here, we studied the role of an ER folding assistant protein Cpr5p on Z-type ${\alpha}_1$-AT folding. Cpr5p was induced > 2-fold in Z-type ${\alpha}_1$-AT-expressing yeast cells compared with the wild type. Knockout of CPR5 exacerbated cytotoxicity of Z-type ${\alpha}_1$-AT, and re-introduction of CPR5 rescued the knockout cells from aggravated cytotoxicity caused by the ${\alpha}_1$-AT variant. Furthermore, Cpr5p co-immunoprecipitated with Z-type ${\alpha}_1$-AT and facilitated its protein folding. Our results suggest that protein-folding diseases may be suppressed by folding assistant proteins at the site of causal protein biosynthesis.

• BMB Reports
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• 제36권3호
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• pp.265-268
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• 2003

Retinoic acid (RA) can transform the Golgi apparatus (GA) into a diffuse vacuolar aggregate and increase the toxicity of some immunotoxins that enter into cells by receptor-mediated endocytosis. An ultramorphological study of the RA-induced GA disruption was performed on F2000 fibroblasts. Cultures were treated with 0.11 to $30\;{\mu}M$ RA for 7 - 180 min. The endocytosis of Limax flavus agglutinin-peroxidase conjugate (LFA), and the interactions between a phorbol ester (PMA) and RA concerning GA disruption, were examined. Exposure to $0.33\;{\mu}M$ RA for 20 min transformed the GA into vacuolar aggregate. These vacuoles were not involved in endocytosis since they remained unstained after endocytosis of LFA. However, the lysosomes were involved in endocytosis, as they were strongly stained. Therefore, a RA-induced shift towards lysosomal routing of the entered LFA was presumed. Exposure to PMA made cells resistant to the Golgi-disturbing effects of RA, indicating that protein kinase C plays an important role in this process.

• EDISON SW 활용 경진대회 논문집
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• 제5회(2016년)
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• pp.55-61
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• 2016

$A{\beta}_{40}$ peptides form oligomers that later aggregate into a plaque, which is deemed to be a leading cause of Alzheimer's Disease. Its non-crystalline morphology has limited an understanding of comprehensive structural study. In this research, computational biomolecular simulations were performed in the following order: solvent and ion addition in a box, energy minimization of protein, equilibration, and periodic boundary condition disaggregation of a monomer from fibril. The result founded the two-fold model is 25% more stable in the simulation environment, and the steric zippers held on most tightly until 220 ps of simulation. The study supports the previous findings that two-fold aggregate $A{\beta}_{40}$ is more stable at 310 K and discusses further how much contribution steric-zipper and hydrogen bonding are making.

• Journal of Plant Biology
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• 제37권2호
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• pp.151-158
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• 1994

Aleurone layers of normal and vp1 mutant maize kernels were extracted and centrifuged at 100,000g to yield a cytosol fraction. Binding of [3H]ABA cis, trans (＋)ABA to a soluble macromolecular components present in the cytosol was demonstrated by Sephadex chromatography and non-denaturing PAGE. The binding component was of high molecular weight and seems to be an aggregate of proteins. A rapid DEAE-cellulose filter method for assaying bound [3H]ABA to a soluble protein was adapted. Binding assays were performed with cytosol that had been preheated or incubated with several enzymes, indicating that heat and protease treatments disrupted the binding. This suggested that binding occurred to proteins. Some properties of the ABA binding proteins were described. The [3H]ABA binding were reduced dramatically when unlabeled ABA was added as a competitor, suggesting a specific binding of [3H]ABA. Gel filtration profiles and autoradiogram of [3H]ABA binding showed no difference in the binding components of Vp1 and vp1/vp1 mutant cytosol, indicating that Vp1 protein is not a sole ABA binding protein.

• 한국자기공명학회논문지
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• 제26권1호
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• pp.17-20
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• 2022

High-pressure (HP) NMR is a versatile tool to investigate diverse features of proteins. This technique has been particularly powerful to elucidate structural dynamics that only populates sufficiently in a pressurized condition. Amyloidogenic proteins, which are prone to aggregate and form amyloid fibrils, often maintains highly dynamic states in its native or aggregation-prone states, and HP NMR contributed much to advance our understandings of the dynamic behaviors of amyloidogenic proteins and the molecular mechanisms of their aggregation. In this mini review, we therefore summarize recent HP NMR studies on amyloid-beta (Aβ), the representative amyloidogenic intrinsically disordered protein (IDP).

• 미생물학회지
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• 제43권3호
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• pp.166-172
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• 2007

ERM 단백질은 23S rRNA의 A2058에 methylation시킴으로써 macrolide-lincosamide- streptogramin B $(MLS_B)$계 항생제의 부착을 저해하여 항생제의 활성을 억제하는 내성 인자 단백질로 monomethylase와 dimethylase로 나누어진다. Dimethylase와 비교되는 monomethylase의 특성을 밝히기 위해 dimethylase (ErmSF)와 monomethylase (TlrD)를 동시에 보유한 Streptomyces fradiae에서 tlrD를 클론하고 대장균에서 최초로 대략생산을 시도하여 $37^{\circ}C$에서 세포내 전체 단백질의 55%를 차지할 정도로 대량생산된 불용성 단백질을 얻어내었다. 그러나 ErmSF와는 달리 낮은 온도에서 대량생산된 단백질이 용해성 단백질로 전환되지 않고 불용성 단백질로 남아있었다. Thioredoxin과 샤페론인 GroESL은 모두 ErmSF의 경우와 마찬가지로 용해성 단백질로의 전환에 도움을 주지 않았다. 이러한 차이점은 천재까지 전혀 밝혀지지 않은 단백질내의 구조적 특성에 의한 monomethylase와 dimethylase의 차이점을 밝힐 수 있다는 가능성을 말해주는 것으로 추정된다. 그러나 ErmSF의 경우와 동일하게 SDS-PAGE에서 검색되지 않은 미량의 발현된 용해성 단백질이 TlrD를 함유한 세포에 항생제에 대한 내성을 나타내게 하였고 이렇게 발현된 내성은 monomethylase에 의한 내성에서 기대되는 내성과 일치하였다.

• Archives of Pharmacal Research
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• 제25권3호
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• pp.381-386
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• 2002

The objective of this study was to link conformational changes of proteins at a water/methylene chloride interface to their destabilization upon emulsification. When 4 aqueous protein solutions (bovine serum albumin, $\beta$-lactoglobulin, ovalbumin, or ribonuclease) were emulsified in methylene chloride, considerable proportions of all the proteins became water insoluble aggregates. There were also noticeable changes in the compositions of their water-soluble species. A series of water/methylene chloride interfacial reactions upon the proteins was considered a major cause of the phenomena observed. Based on this supposition, the interfacial tension was determined by a Kruss DVT-10 drop volume tensiometer under various experimental conditions. It substantiated that the interfacial tension was high enough to cause the adsorption of all the proteins. Under our experimental conditions, their presence in the aqueous phase resulted in reductions of the interfacial tension by the degrees of 8.5 - 17.1 mN $m^{-1}$. In addition, dynamic changes in the interfacial tension were monitored to compare relative rates at which the adsorbed proteins underwent conformational, structural rearrangements at the interface. Such information helped make a prediction about how easily proteins would denature and aggregate during emulsification. Our study indicated that emulsifying aqueous protein solutions in organic solvents should be handled with care, due to adverse interfacial effects.

• 한국생물공학회:학술대회논문집
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• 한국생물공학회 2000년도 춘계학술발표대회
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• pp.541-545
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• 2000

For the sustained release formulation of recombinant human growth hormone (rhGH), dissociable rhGH aggregates were microencapsulated within poly(D,L-lactic-co-glycolic acid) [PLGA] microparticles. rhGH aggregates with 2 - 3 m Particle diameter were first produced by adding a small volume of aqueous rhGH solution into a partially water miscible organic solvent phase(ethyl acetate) containing PLGA. These rhGH aggregates were then microencapsulated within PLGA polymer phase by extracting ethyl acetate into an aqueous phase pre-saturated with ethyl acetate. The resultant microparticles were 2 - 3 m in diameter similar to the size of rhGH aggregates, suggesting that PLGA polymer was coated around the protein aggregates. Release profiles of rhGH from these microparticles were greatly affected by changing the volume of the incubation medium. The release rhGH species consisted of mostly monomeric form with having a correct conformation. This study reveals that sustained rhGH release could be achieved by microencapsulating reversibly dissociable protein aggregates within biodegradable polymers.

• 한국자기공명학회논문지
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• 제19권2호
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• pp.74-82
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• 2015

Ryanodine receptor (RyR) is one of the two major $Ca^{2+}$ channels in membranes of intracellular $Ca^{2+}$ stores and is found in sarcoplasmic reticulum (SR), endoplasmic reticulum (ER). RyR1 is also the major calmodulin-binding protein of sarcoplasmic reticulum membranes. Residues 4064-4210 in the RyR1 polypeptide chain has similar primary sequence with calmodulin (CaM) and was designated as CaM-like domain (CaMLD). When expressed as a recombinant peptide, CaMLD showed several CaM-like properties in previous studies. Still, previous studies of CaMLD were focused on protein-protein interactions rather than its own properties. Here, we studied the expression of CaMLD and its sub-domains corresponding to each lobe of CaM in Escherichia coli. CaMLD could be obtained only as inclusion body, and it was refolded using urea solubilization followed by dialysis. Using spectroscopic approaches, such as NMR, circular dichroism, and gel filtration experiment, we found that the refolded CaMLD exists as nonspecific aggregate, even though it has alpha helical secondary structure. In comparison, the first half of CaMLD (R4061-4141) could be obtained as natively soluble protein with thioredoxin fusion. After the removal of the fusion tag, it exhibited folded and helical properties as shown by NMR and circular dichroism experiments. Its oligomeric status was different from CaMLD, existing as dimeric form in solution. However, the second half of the protein could not be obtained as soluble protein regardless of fusion tag. Based on these results, we believe that CaMLD, although similar to CaM in sequence, has quite different physicochemical properties and that the second half of the protein renders it the aggregative properties.