• 대한수의학회지
• /
• 제37권2호
• /
• pp.367-374
• /
• 1997

Bovine rotaviruses(A, 288, 55086 strains) isolated from fecal samples in Korea were propagated onto MA104 cells and were confirmed tentatively as G6, G8, and G10, respectively, by RFLP analysis. Full-length VP7 gene of these isolates was amplified by reverse transcriptase polymerase chain reaction(RT-PCR) using VP7 specific primers and cloned into TA vector. Nucleotide and deduced amino acid sequences of VP7 genes of the isolates were determined and compared with those of bovine rotavirus reference strains(NCDV; G6, UK; G6, Cody I-801; G8 and B223; G10). A, 288 and 55086 isolates showed high degree of nucleotide sequence homology with NCDV and UK(93% and 94%), Cody I-801(86%) and B223(97%), respectively, However, they showed 71~74% of nucleotide sequence homlogy with bovine rotavirus reference strains which belong to different serotypes. From the results of deduced amino acid sequence homology analysis, three isolates showed 94~96% of homology with the same serotype reference strains but 80~84% of homology with the different serotype reference strains. Three bovine rotavirus isolates, A, 288 and 55086 strains, were confirmed as G6, G8, and G10, respectively, by nucleotide and deduced amino acid sequence analysis.

• 한국버섯학회지
• /
• 제17권4호
• /
• pp.185-190
• /
• 2019

리그닌 분해 담자균류로 알려져 있는 느타리버섯균 No.42는 망간퍼옥시데이즈(MnP) 및 락게이즈(Lac)를 생산하였으나 글루코오스-펩톤-이스트-밀기울(GPYW)배지를 이용한 정치배양조건하에서 리그닌퍼옥시데이즈(LiP)활성은 검출되지 않았다. 한편, 동일배지에서 망간퍼옥시데이즈 활성은 11일째 80(3.6) U/flask(ml)의 최대 생산되었다. 망간퍼옥시데이즈 분리정제는 Sepha-ros CL-6B 및 Mono-Q 컬럼순으로 수행하였으며 주요 망간퍼옥시데이즈 isozyme은 단일밴드로 분자량은 36.4KDa이였다. N-말단으로부터 19개의 아미노산 배열은 단백질 자동 분석 장치로 분석한 결과 ATCADGRTTANAACCVLFP를 나타내었다. 느타리버섯균 No.42의 정치배양조건 하에서 세포외로 생산되는 중요한 망간퍼옥시데이즈 isozyme의 N-말단 아미노산 배열은 이전에 보고된 MnP3의 아미노산 배열과 동일하였다.

• 한국자원식물학회지
• /
• 제16권3호
• /
• pp.238-244
• /
• 2003

더덕 뿌리에서 유래한 EST 라이브러리로부터 dehydrin 유전자와 높은 상동성 을 나타내는 full clone cDNA를 얻었다. 더덕의 dehydrin, ClDhn1은 893 bp의 cDNA로 159개의 아미노산을 코딩하는 480 bp의 ORF를 가지고 있다. ClDhn1의 아미노산을 분석해 보면, 전체적으로 높은 친수성을 나타내며, lysine이 풍부한 K 반복구간(KIKEKLPG)을 카르보닐기 쪽에 2개 가지고 있다. 또한, 여러 dehydrin들의 공통적인 특징인 7개의 연속적인 serine잔기가 첫 번째 K반복 구간 앞에 위치한다. 그러나, 아미노기쪽의 DEYGNP보존 구간은 변형(DEHGNP)되어 있다. ClDhn1 유전자는 전사 단계에서 더덕의 뿌리에서 가장 높은 발현 양상을 보이며, 줄기와 잎에서는 적은 양이 발현되었다.

• The Plant Pathology Journal
• /
• 제19권3호
• /
• pp.171-176
• /
• 2003

The complete nucleotide sequences of the genomic RNAs of Soybean mosaic virus strains GS (SMV-G5) and G7H (SMV-G7H) were determined and compared with sequences of other SMV strains. Each viral RNA was determined to be 9588 nucleotides in length excluding the poly (A) tail and contained an open reading frame to encode a polyprotein subsequently processed into up to ten proteins by proteolytic cleavage. Com-parison of the amino acid sequences with those of other SMV strains showed high percentage of amino acid sequence homology with the same genome organization. The nucleotide and the deduced amino acid sequences between SMV-G5 and SMV-G7H were greater than 99% identity. When compared with those of other SMV strains in a phylogenetic analysis of the nucleotide and deduced amino acid sequences, they formed a distinct virus clade showing over 97％ amino acid identity, but were more distantly related to the other potyvirus (44.1-69.6% identity). Interestingly, SMV G7H strain caused a severe mosaic or necrosis symptom in soybean cultivars including Jinpum-1, Jinpum-2, and Sodam, whereas, no symptom was observed in SMV-G5 inoculation. Complete nucleotide sequences of these strains will give clues for determining symptom determinant(s) in future research.

• Asian-Australasian Journal of Animal Sciences
• /
• 제23권2호
• /
• pp.272-278
• /
• 2010

In this study, we cloned, sequenced and characterized porcine y+L Amino Acid Transporter-1 (y+LAT1). By screening a translated EST database with the protein sequence of the human $y^{+}$LAT1 and by using rapid amplification of cDNA ends (RACE), the full-length cDNA encoding porcine $y^{+}$LAT1 was isolated from porcine intestine RNA. It was 2,111 bp long, encoding a 511 amino acid trans-membrane glycoprotein composed of 12 transmembrane domains. The predicted amino acid sequence was found to be 91%, 90%, 87% and 87% identical to those of cattle, human, mouse and rat $y^{+}$LAT1 respectively. Real-time RT-PCR results indicated that the small intestine had the highest $y^{+}$LAT1 mRNA abundance and the lung had the lowest $y^{+}$LAT1 mRNA abundance. Baby hamster kidney (BHK) cells transfected with green fluorescent protein (GFP) tagged porcine $y^{+}$LAT1 cDNA indicated that the cellular localization of the gene product in BHK was on the plasma membrane.

• Journal of the Korean Association of Oral and Maxillofacial Surgeons
• /
• 제30권3호
• /
• pp.186-192
• /
• 2004

Purpose: Human tooth proteins are highly heterogeneous, comprising diverse proteins derived from a number of genes. The attempts to identify protein for activity of tooth matrix proteins have been defied by several factors. First, the amount of proteins within teeth is very small relative to many extracellular matrix proteins of other tissues. Second, the bioassay system is tedious and needed for long time. Therefore we tried to find easy techniques, which increase the product rate, and an assay of small proteins, with which amino acid sequence is possible without additional procedures. Materials and Methods: Total protein were extracted from 300 g enamel removed teeth and 600 g teeth with 4 mol/L guanidine HCl and purified by gel chromatography. Aliquot of proteins was implanted into muscle pouches in Sprague-Dawley rats for bioassay. By SDS-PAGE and membrane blotting, molecular weight of each protein was estimated and a partial amino acid sequence was obtained. Each fraction blotted on the membrane was cut out and inserted in rat ectopic model. Results: In dissociative method, total tooth proteins were obtained 1mg/ml from enamel removed teeth and 3.5 mg/ml from teeth. In SDS-PAGE, four clear bands at the sites corresponding to 66, 40, 20 and 18 kD. Especially The 66 kD band was clearly exhibited. Amino acid sequencing from tooth could be possible using PVDF membrane blotting technique. In amino acid sequencing, 66 kD protein was identified as albumin. Conclusion: Compared with conventional method for extraction of teeth protein and bioassay of proteins, the methods in this study were easy, time-saving and more productive technique. The matured tooth proteins omitting additional procedure of mechanical removal of enamel were simply analyzed using blotted PVDF membrane. This method seems to make a contribution as a technique for bioassay and amino acid sequencing of protein.

• International Journal of Industrial Entomology and Biomaterials
• /
• 제23권2호
• /
• pp.231-238
• /
• 2011

Attacin is an insect antibacterial protein that plays an important role in immune response to injury and infection. In this report, we have isolated and characterized of cDNA encoding for the attacin from the immunized larvae of swallowtail butterfly, $Papilio$ $xuthus$. A full length cDNA of $P.$ $xuthus$ attacin was obtained by employing annealing control primer (ACP)-based differential display PCR and 5' RACE. The complete $P.$ $xuthus$ attacin cDNA was comprised of 949 bp encoding a 250 amino acid precursor. It contains a putative 18 amino acid signal peptide sequence, a 42 amino acid propeptide sequence, and a 190 amino acid mature protein with a theoretical molecular mass of 19904.01 and a pI of 9.13. The putative mature protein of $P.$ $xuthus$ attacin showed 48-52% and 24-30% identity in amino acid sequences with that of lepidopteran and dipteran insects, respectively. Semiquantitive RT-PCR results revealed that the transcript of $P.$ $xuthus$ attacin gene was up-regulated at significant levels after injection with bacterial lipopolysaccharide (LPS). We sub-cloned cDNA fragment encoding mature $P.$ $xuthus$ attacin into the expression vector, highly expressed in $E.$ $coli$ BL21 cells, and its antibacterial activity was analyzed. Recombinant $P.$ $xuthus$ attacin evidenced considerably antibacterial activity against Gram-negative bacteria, $E.$ $coli$ ML 35 and $Klebsiella$ $pneumonia$.

• 미생물학회지
• /
• 제39권1호
• /
• pp.1-7
• /
• 2003

단일 탄소원과 질소원 및 에너지원으로 aniline을 이용하는 Delftia sp. JK-2에서 분리 정제한 catechol 2,3-dioxygenase (C2,3O)의 특성과 N-말단 아미노산 및 DNA 서열을 분석하였다. C2,3O의 특성을 조사하기 위하여 aniline에서 배양한 Delftia sp. JK-2를 초음파 분쇄기로 파쇄하였으며, ammonium sulfate precipitation과 DEAE-sepharose로 정제하였다. 정제된 C2,3O의 고유활성(specific activity)은 약4.72 unit/mg이었으며, C2,3O는 catechol과4-methylcatechol 대해서 효소활성을 나타내었다. C2,3O는$30^{\circ}C$와pH 8.0에서 최적 활성을 나타내는 것으로 조사되었으며, $Ag^{+}$, $Hg^{+}$,그리고 $Cu^{2+}$는 Deftia sp. JK-2의 C2,3O 활성을 억제하였다. SDS-PAGE에 의해 측정 된C2,3O의 분자량은 약 35 KDa이었으며, N-말단 아미노산 서열을 분석한 결과, $^{1}MGVMRIG-HASLKVMDMDA- AVRHYENV^{26}$로 확인되었다. 이 N-말단 아미노산 서열은 Pseudomonas sp. AW-2와 Coma-monas sp. Js765의 C2,30와 일치하는 것으로 나타났으며, 얻어진 결과를 토대로 primer를 제작하여 polymerase chain reaction (PCR)을 실시하였다. PCR을 통해 얻어진 Delftia sp. JK-2의 C2,30유전자 DNA서열을 분석하여 상동성 조사를 하였다. DNA서열의 상동성 조사는 유추되는 아미노산 서 열로 바꾸어서 실시하였으며,그 결과 Deftia JK-2의 C2,3O는Pseudomonas sp. AW-2의 C2,3O(100%)와 Coma-monas sp. JS76S의 C2,3O(97%)에서 높은 상동성 이 확인되었다.

• Parasites, Hosts and Diseases
• /
• 제44권3호
• /
• pp.251-254
• /
• 2006

Monoclonal antibody (mAb) Tg786 against Toxoplasma gondii has been found to detect a 42-kDa rhoptry protein (ROP6) which showed protease activity and host cell binding characteristics after secretion. Using the mAb, a colony containing a 3'-UTR was probed in a T. gondii cDNA expression library. A full length cDNA sequence of the rhoptry protein was completed after 5'-RACE, which consisted of 1,908 bp with a 1,443 bp ORF. The deduced amino acid sequence of ROP6 consisted of a polypeptide of 480 amino acids without significant homology to any other known proteins. This sequence contains an amino terminal stop transfer sequence downstream of a short neutral sequence, hydrophilic middle sequence, and hydrophobic carboxy terminus. It is suggested that the ROP6 is inserted into the rhoptry membrane with both N- and C-termini.

• Fisheries and Aquatic Sciences
• /
• 제24권4호
• /
• pp.163-170
• /
• 2021

Peptides are naturally occurring biological molecules that are found in all living organisms. These biologically active peptides play a key role in various biological processes. The aim of this study is the extraction and the purification of bioactive materials that induce relaxation of an apical muscle from the pyloric caeca of Patiria pectinifera. The acidified pyloric caeca extract was partially separated by the solid phase extraction using a stepwise gradient on Sep-Pak C18 cartridge. Among the fractions, materials eluted with 60% methanol/0.1% trifluoroacetic acid was put a thorough of a series of high performance liquid chromatography (HPLC) steps to isolate a neuropeptide with relaxation activity. The purified compound was eluted at 28% acetonitrile in 0.1% trifluoroacetic acid with retention time of 25.8 min on the CAPCELL-PAK C18 reversed-phase column. To determine the molecular weight and the amino acid sequence of the purified peptide, LC-MS and Edman degradation method were used, respectively. The primary structure of the peptide was determined to be FGMGGAYDPLSAGFTD which corresponded to the amino acid sequence of a starfish myorelaxant peptide (SMP) isotype (SMPb) found in the cDNA sequence encoding SMPa and its isotypes. In this study, a muscle relaxant neuropeptide (SMPb) has been isolated from pyloric caeca of starfish P. pectinifera. This is the first report of SMPb isolation on the protein level from P. pectinifera.