• International Journal of Industrial Entomology and Biomaterials
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• 제17권2호
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• pp.223-228
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• 2008

A novel beetle antimicrobial protein from stimulated Copris tripartitus and the corresponding gene were isolated in parallel through differential display-PCR and expression in Escherichia coli. To find cDNA clones responsible for bacteria resistance, the suppression subtractive hybridization and GeneFishing differentially expressed genes system were employed in the dung beetle, Copris tripartitus immunized with lipopolysaccaride. One cDNA clone from eight subtracted clones was selected through dot blot analysis and confirmed by northern blot analysis. The 516-bp, selected cDNA clone was determined by 5' and 3' rapid amplication of cDNA ends and cloned into the GST fusion expression vector pGEX-4T-1 for expression of the protein. The expressed protein was predicted 14.7 kDa and inhibited the growth of gram-negative bacteria such as Escherichia coli and Pseudomonas aeruginosa. These results implied that the expressed protein is related to immune defense mechanism against microorganism.

• BMB Reports
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• 제34권5호
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• pp.428-433
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• 2001

A defect in uvsC of Aspergillus nidulans caused high methyl methansulfonate (MMS)-sensitivity, hyporecombination, and a lack of UV induced mutation. The uvsC gene of Aspergillus nidulans shares a sequence similarity with the RAD51 gene of Saccharomyces cerevisiae. In this study, in vitro and in vivo tests were conducted in order to determine whether or not the UVSC protein had functional similarities to RAD51, the recombination enzyme in yeast. The purified recombinant UVSC protein, following expression in Escherichia coli, showed binding activity to single-stranded DNA (ssDNA), when both ATP and magnesium are present. In addition, ATPase activity was also demonstrated and its activity was stimulated in the presence of ssDNA. The UVSC protein that was expressed under the ADH promoter in S. cerevisiae suppressed in part the sensitivity to MMS of the rad51 null mutant. Similarly, when the uvsC cDNA was expressed from the nmt promoter, the MMS sensitivity of the rhp51 null mutant of Schizosaccharomyces pombe was partially complemented. These results indicate that the A. nidulans UVSC protein is a functional homologue of the RAD51 protein.

• 센서학회지
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• 제14권3호
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• pp.137-143
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• 2005

Research in the field of biosensor has enormously increased over the recent years. The metal-oxide semiconductor field effect transistor (MOSFET) type protein sensor offers a lot of potential advantages such as small size and weight, the possibility of automatic packaging at wafer level, on-chip integration of biosensor arrays, and the label-free molecular detection. We fabricated MOSFET protein sensor and proposed the gold-black electrode as the gate metal to improve the response. The experimental results showed that the output voltage of MOSFET protein sensor was varied by concentration of albumin proteins and the gold-black gate increased the response up to maximum 13 % because it has the larger surface area than that of planar-gold gate. It means that the expanded gate allows a larger number of ligands on same area, and makes the more albumin proteins adsorbed on gate receptor.

• International Journal of Industrial Entomology and Biomaterials
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• 제1권2호
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• pp.171-175
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• 2000

Determined sequences of 384 randomly selected clones in a PCR-based cDNA library of Antheraea yamamai could identify expressed sequence tags (ESTs) of highly expressed gene. One EST (fibroin) appeared 15 times, one EST (40S ribosomal protein S18) twelve times, one EST (ribosomal protein S24a) eleven times, ten times (ribosomal protein S8), nine times (60S ribosomal protein L10A), seven times (60S ribosomal protein S15A, S17, S17 and seroin), six times (ribosomal protein S8), five times (ribosomal protein S24, mariner transposase and P8 protein), four times (serpin 2), three times (heat shock protein 70 and poly A binding protein), and the remaining 6 ESTs twice (amylase, KIAA1006, elongation factor-1, transposon mag, translation initiation factor 4C, QM protein, transposase). Therefore, the 94 EST make it possible to identify 24 redundant clones that are candidates for highly expressed genes in posterior silk gland of this insect. The 24 redundant EST clones were identified in GenBank, but none of them was related to A. yamamai, suggesting that there are many unidentified genes which are highly expressed in the A. yamamai genome.

• Journal of Applied Biological Chemistry
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• 제44권3호
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• pp.119-124
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• 2001

NTR1 gene of Brassica campestris L. ssp. perkinensis encodes a floral nectary-specific methyltransferase. In this study, the NTR1 cDNA was expressed in E. coli to examine the enzymatic characteristics of the protein product. The GST-NTR1 fusion protein was purified to near homogeneity, showing that the size of NTR1 was 44 kDa. The protein reacted specifically with jasmonic acid (JA), consuming methyl group from S-adenosyl-L-methionine (SAM). GC-MS analysis revealed that the compound produced was authentic methyl jasmonate (MeJA), suggesting that NTR1 is an S-adenosyl-L-methionine: jasmonic acid carboxyl methyltransferase. Km values of NTR1 for JA and SAM were 38.0 and $6.4{\mu}M$, respectively. Optimal activity of the NTR1 was observed at $20^{\circ}C$, pH 7.5, in the presence of 100-150 mM KCl. Thus, kinetic properties, thermal characteristics, optimal pH, and ion-dependency of the NTR1 activity were almost identical to those of Arabidopsis JA methyltransferase JMT, indicating that these two proteins are orthologues of each other.

• Molecules and Cells
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• 제38권3호
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• pp.243-250
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• 2015

Patterning of the polar axis during the early leaf developmental stage is established by cell-to-cell communication between the shoot apical meristem (SAM) and the leaf primordia. In a previous study, we showed that the DRL1 gene, which encodes a homolog of the Elongator-associated protein KTI12 of yeast, acts as a positive regulator of adaxial leaf patterning and shoot meristem activity. To determine the evolutionally conserved functions of DRL1, we performed a comparison of the deduced amino acid sequence of DRL1 and its yeast homolog, KTI12, and found that while overall homology was low, well-conserved domains were presented. DRL1 contained two conserved plant-specific domains. Expression of the DRL1 gene in a yeast KTI12-deficient yeast mutant suppressed the growth retardation phenotype, but did not rescue the caffeine sensitivity, indicating that the role of Arabidopsis Elongator-associated protein is partially conserved with yeast KTI12, but may have changed between yeast and plants in response to caffeine during the course of evolution. In addition, elevated expression of DRL1 gene triggered zymocin sensitivity, while overexpression of KTI12 maintained zymocin resistance, indicating that the function of Arabidopsis DRL1 may not overlap with yeast KTI12 with regards to toxin sensitivity. In this study, expression analysis showed that class-I KNOX genes were downregulated in the shoot apex, and that YAB and KAN were upregulated in leaves of the Arabidopsis drl1- 101 mutant. Our results provide insight into the communication network between the SAM and leaf primordia required for the establishment of leaf polarity by mediating histone acetylation or through other mechanisms.

• 식물분류학회지
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• 제38권1호
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• pp.51-61
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• 2008

잎은 무한생장기관으로 잎의 극성제어에 많은 유전적인 요소가 필요하다. 이들 극성은 잎의 초기발생과정에서 제어되기 시작하고, 정단분열조직과 잎기관의 원기와의 제어를 담당하는 인자들에 의해서 결정이 된다. 본 연구에서는 가늘고 바늘처럼 생긴 잎을 가진 deformed root and leaf1 (drl1) 돌연변이체를 유전학적 해석하였고, 그 결과 DRL1 유전자는 정단분열조직과 잎의 극성축을 제어하고 있는 것으로 판명되었다. 이 DRL1 유전자는 효모의 KTI12 유전자 산물과 유사한 단백질인 Elongator associate protein을 만들어 내는 것으로 판명되었다. 또한, 이 단백질의 아미노산 서열이 원핵생물에서부터 진핵생물까지 광범위하게 진화적으로 보존되고 있는 것으로 밝혀졌다. 특히, DRL1 단백질과 유사한 식물의 단백질은 계통해석 결과 단일계통을 나타내고 있는 것으로 나타났고, 이는 이 단백질들이 육상식물의 진화과정에서 잘 보존되고 있음을 시사하고 있다.

• BMB Reports
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• 제35권5호
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• pp.498-507
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• 2002

A gene that encodes a thermostable protease, coined thermicin, has been isolated from Thermoanaerobacter yonseiensis that is expressed and characterized in E. coli.. In order to elucidate the molecular characteristics on thermostability of the enzyme, molecular modeling and mutagenesis technology were applied. In the modeling structure, the structural core, including the active site, was well conserved; whereas, the two loop regions were unique when compared to thermitase. The mutant enzyme with the small loop deleted (D190-I196), based on modeling structural information, showed identical enzyme activity. However, when the large loop was deleted (P233-P244), a little lower $K_m$ and even a lower kcat was found. This indicates that the large loop could influence catalytic activity. However, the unfolding temperature ($T_m$), which was determined by a differential-scanning calorimetry for the mutant enzyme deleted the small loop, was $96^{\circ}C$. This is $14^{\circ}C$ lower than that for the parent thermicin. These results suggest that the small loop may play a role in maintaining the proper folding of the enzyme at high temperatures, whereas the large loop might be related to catalysis.

• International Journal of Industrial Entomology and Biomaterials
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• 제17권2호
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• pp.217-221
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• 2008

Dung beetle larvae at the $3^{rd}$ instar were injected with lipopolysaccaride and inducible proteins were examined within a pI level of 3-10 and a size level by proteomics, including 1-D SDS PAGE analysis and antibacterial assay. The immune infected larvae extracts provided seven protein bands in one-dimensional electrophoresis and its antibacterial activity also checked. Hemolymph protein from immune infected larvae of the dung beetle were separated by twodimensional gel electrophoresis and compared with those from native larvae. In 2-D gel electrophoresis, we detected 63 immune infected unique and 32 up-regulated proteins, and 36 proteins that were down-regulated or not present in treated gel. Ten protein spots from unique proteins and those presented as different level of abundance in infected and native larvae were specially expressed. These differentially expressed proteins were proposed to be involved in the defense mechanism against microorganism.

• 한국응용과학기술학회지
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• 제19권2호
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• pp.79-85
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• 2002

The cutaneous tolerability of detergent formulations can be improved by means of suitable additives. They complex the surfactant molecules lowering the concentration of their free monomeric species. Proteins derivatives used as additives for detergency are usually prepared by partial hydrolysis of plant reserve proteins. The main purpose of the hydrolytic cleavage is to make them water soluble and suitable for liquid products. Water solubility and stability are obtained by means of complexation with surfactants which also increase their actual hydrophobicity, an important parameter affecting cosmetic properties of proteins. Transepidermal water loss (TEWL) and electric capacitance (EC) have been adopted as investigation technigues to evaluate the skin integrity/damage in vitro tests, The performance of native wheat protein / surfactant complexes has been compared with traditional protein hydrolysates as detergent additives. The results show a noticeable reduction of skin irritation in surfactant formulations with addition of native wheat protein.