• Development and Reproduction
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• v.20 no.4
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• pp.297-304
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• 2016

Lectins belong to the pattern-recognition receptors (PRRs) class and play important roles in the recognition and elimination of pathogens via the innate immune system. Recently, it was reported that lily-type lectin-1 is involved when a pathogen attacks in the early immune response of fish. However, this study is limited to information that the lectin is involved in the innate immune response against viral infection. In the present study, the lily-type lectin-2 and -3 of Oplegnathus fasciatus (OfLTL-2 and 3) have been presented to be included B-lectin domain and two D-mannose binding sites in the amino acid sequence that an important feature for the fundamental structure. To investigate the functional properties of OfLTLs, the tissue distribution in the healthy rock bream and temporal expression during early developmental stage analysis are performed using quantitative real-time PCR. OfLTL-2 and 3 are predominantly expressed in the liver and skin, but rarely expressed in other organ. Also, the transcripts of OfLTLs are not expressed during the early developmental stage but its transcripts are increased after immune-related organs which are fully formed. In the challenge experiment with RBIV (rock bream iridovirus), the expression of OfLTLs was increased much more strongly in the late response than the early, unlike previously known. These results suggest that OfLTLs are specifically expressed in the immune-related tissues when those organs are fully formed and it can be inferred that the more intensively involved in the second half to the virus infection.

• Korean Journal of Veterinary Research
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• v.37 no.3
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• pp.541-546
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• 1997

There are two conserved regions with a significantly high amino acid sequence homology among the A subunits of STX, SLTs and ricin. To produce an inactive Verotoxin-2 (VT-2), two different mutants, pE167D and pDE5A, were constructed by site-directed mutagenesis, respectively, on the basis of the previous reports that two regions lie within the active-site clefts of the A subunits of ricin and STX family. The cytotoxicity ($10^3$ $CD_{50}/ml$) of VT-2 holotoxin with E167D mutation was reduced by $10^3$-fold compared with wild-type level. In addition, VT-2 with DE5A ($Trp_{202}GlyArgIleSer_{206}$) deletion mutation showed a significantly low cytotoxicity ($10^1$ $CD_{50}/ml$), resulting in $10^5$- and $10^2$-fold reductions, respectively, compared with the wild-type and E167D mutatant. SDS-PAGE for protein samples showed a 33-kDa band corresponding to the A subunit of VT-2. These results indicate that reduction in cytotoxic activity was affected not by amount of VT-2 protein produced but by mutation.

• Horticultural Science & Technology
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• v.31 no.6
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• pp.748-755
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• 2013

This study was conducted to find a salt tolerance gene in Brassica rapa. In order to meet this objective, we analyzed data from a KBGP-24K oligo chip [BrEMD (Brassica rapa EST and microarray database)] of the B. rapa ssp. pekinensis 'Chiifu' under salt stress (250 mM NaCl). From the B. rapa KBGP-24K microarray chip analysis, 202 salt-responsive unigenes were primarily selected under salt stress. Of these, a gene with unknown function but known full-length sequence was chosen to closely investigate the gene function. The selected gene was named BrSSR (B. rapa salt stress resistance). BrSSR contains a 285 bp open reading frame encoding a putative 94-amino acid protein, and a DUF581 domain. The pSL94 vector was designed to over-express BrSSR, and was used to transform tobacco plants for salt tolerance analysis. T1 transgenic tobacco plants that over-expressed BrSSR were selected by PCR and DNA blot analyses. Quantitative real-time RT PCR revealed that the expression of BrSSR in transgenic tobacco plants increased by approximately 3.8-fold. Similar results were obtained by RNA blot analysis. Phenotypic characteristics analysis showed that transgenic tobacco plants with over-expressed BrSSR were more salt-tolerant than the wild type control under 250 mM NaCl for 5 days. Based on these results, we hypothesized that the over-expression of BrSSR may be closely related to the enhancement of salt tolerance.

• Journal of Life Science
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• v.22 no.8
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• pp.1024-1033
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• 2012

A gene encoding a novel geranylgeranyl pyrophosphate (GGPP) synthase from Kocuria gwangalliensis has been cloned and expressed in Escherichia coli. The deduced amino acid sequence showed 59.6% identity with a putative GGPP synthase (CrtE) from K. rhizophila. An expression plasmid containing the crtE gene was constructed, and E. coli cells containing this plasmid produced a recombinant protein with a theoretical molecular mass of 41 kDa, corresponding to the molecular weight of GGPP synthase. Due to the lack of crtE, crtB, and crtI in E. coli, the biosynthesis of lycopene was only obtained when the plasmid pCcrtE was co-transformed into E. coli expressing the pRScrtBI-carrying carotenoid biosynthesis crtB and crtI genes, which were sub-cloned from Paracoccus haeundaensis. The biochemical studies on the expressed proteins were performed via HPLC. The results obtained from this study will provide a wider base of knowledge regarding the primary structure of CrtE cloned from K. gwangalliensis at the molecular level.

• Journal of The Korean Society of Inherited Metabolic disease
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• v.14 no.2
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• pp.186-190
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• 2014

Citrullinemia type 2 (citrin deficiency) is an autosomal recessive inborn error metabolism, caused by the SLC25A13 gene mutation. Citrin deficiency is associated with two clinical phenotype; neonatal-onset type II citrullinemia (CTLN2), also known as neonatal intraphepatic cholestasis caused by citrin deficiency (NICCD) and adult-onset CTLN2. Clinical manifestations of NICCD include poor growth, intrahepatic cholestasis, liver dysfunction and increased plasma citrulline, methionine, threonine, arginine. The molecular diagnosis could be confirmed by SLC25A13 gene mutation analysis. A 3-month-old male infant with persistent jaundice was referred for evaluation. Newborn screening was normal at birth. Mild elevation of serum ammonia and AST/ALT were observed. Plasma amino acid analysis showed significantly elevated citrulline, methionine, threonine. DNA sequence analysis of the SLC25A13 gene revealed two compound heterozygous mutations, c.[852_855del]($p.Met285Profs^*2$) and [1180+1G>A]. We suggest that NICCD should be considered as one of the cause of in infants with cholestatic jaundice, although the newborn screening was normal.

• Korean Journal of Microbiology
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• v.37 no.4
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• pp.245-252
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• 2001

Erm proteins, MLS (macrolide-lincosamide-streptogramin B) resistance factor proteins, show high degree of amino acid sequence homology and comprise of a group of structurally homologous N-methyltransferases. On the basis of the recently determined structures of ErmC` and ErmAM, ErmSF was divided into two domains, N-terminal end catalytic domain and C-terminal end substrate binding domain and attempted to overexpress catalytic domain in E. coli using various pET expression systems. Three DNA fragments were used to express the catalytic domain: DNA fragment 1 encoding Met 1 through Glu 186, DNA fragment 2 encoding Arg 60 to Glu 186 and DNA fragment 3 encoding Arg 60 through Arg 240. Among the pET expression vectors used, pET 19b successfully expressed the DNA fragment 3 and pET23b succeeded in expression of DNA fragment 1 and 2. But the overexpressed catalytic domains existed as inclusion body, a insoluble aggregate. To assist the soluble expression of ErmSF catalytic domains, Coexpression of chaperone GroESL or Thioredoxin and lowering the incubation temperature to $22^{\circ}C$ were attempted, as did in the soluble expression of the whole ErmSF protein. Both strategies did not seem to be helpful. Solubilization with guanidine-HCl and renaturation with gradual removal of denaturant and partial digestion of overexpressed whole ErmSF protein (expressed to the level of 126 mg/ι culture as a soluble protein) with proteinase K, nonspecific proteinase are under way.

• Journal of Life Science
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• v.14 no.4
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• pp.636-643
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• 2004

D-Xylose isomerase produced by Lactococcus sp. JK-8, isolated from kimchi, was purified 17-fold of homogeneity, and its physicochemical properties were determined. Although the N-terminal amino acid sequence of D-xylose isomerase was analysed to Ala-Tyr-Phe-Asn-Asp-Ile-Ala-Pro-Ile-Lys, it was not similar to that of Lactobacillus enzyme. The molecular weight of the purified enzyme was estimated to be 180 kDa by gel filtration, 45 kDa by SDS-PAGE and the enzyme was homotetramer. The optimum pH of the enzyme was around 7 and stable between pH 6 and 8. The optimum reaction temperature was 7$0^{\circ}C$ and stable up to 7$0^{\circ}C$ in the presence of 1 mM $Mn^{2+}$. Like other D-xylose isomerases, this enzyme required divalent cation, such as $Mg^{2+}$, $Co^{2+}$, or $Mn^{2+}$ for the activity and thermostability. $Mn^{2+}$was the best activator. Substrate specificity studies showed that this enzyme was highly active on D-xylose. The enzyme had an isoelectric point of 4.8, and fm values for D-xylose was 5.9 mM.

• Journal of KIISE:Databases
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• v.34 no.5
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• pp.396-408
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• 2007

After the Human Genome Project finished the sequencing of a human DNA sequence, the concerns on protein functions are increasing. Since the structures of proteins are conserved in divergent evolution, their functions are determined by their structures rather than by their amino acid sequences. Therefore, if similarities between two protein structures are observed, we could expect them to have common biological functions. So far, a lot of researches on protein structure alignment have been performed. However, most of them use RMSD(Root Mean Square Deviation) as a similarity measure with which it is hard to judge the similarity level of two protein structures intuitively. In addition, they retrieve only one result having the highest alignment score with which it is hard to satisfy various users of different purpose. To overcome these limitations, we propose a novel protein structure alignment algorithm based on MRPD(Maximum of Residue Pair Distance) and SG (Similarity Graph). MRPD is more intuitive similarity measure by which fast tittering of unpromising pairs of protein pairs is possible, and SG is a compact representation method for multiple alignment results with which users can choose the most plausible one among various users' needs by providing multiple alignment results without compromising the time to align protein structures.

• Journal of Life Science
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• v.27 no.11
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• pp.1233-1244
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• 2017

CDPKs have pivotal roles in plant $Ca^{2+}$-mediated transduction signaling. A total of 29 CDPK genes have been identified in rice (Oryza sativa L.), but their key functions have not been completely noted. This study focused on the OsCPK11 gene, which has not been studied, to determine its functional characteristics. A study of tissue-specific expressions revealed that the OsCPK11 gene is expressed in young leaves, mature leaves and flowers of rice. An expression of the gene was also confirmed in gibberellin-treated aleurone layers of rice. Regarding the phenotypic characteristics of Tos17-inserted OsCPK11 mutants, the heights of the mutants were not distinguishable from the heights of wild type plants, but the number of caryopses and the caryopses' weights were significantly statistically different. In addition, many grains of the mutants had white belly materials in their endosperm. The cDNA of the OsCPK11 was cloned, and an OsCPK11 protein of about 60.5 kD was obtained by using a GST affinity chromatography and an SDS-PAGE. An analysis of the amino-acid sequence of the protein indicated that the OsCPK11 protein has the structural characteristics of typical CDPKs. The results provided useful information about the functions of the OsCPK11 gene and further noted the roles CDPKs have in $Ca^{2+}$-mediated signaling in plants.

• KSBB Journal
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• v.24 no.3
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• pp.259-266
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• 2009

AfsR2 in Streptomyces lividans, a 63-amino acid protein with limited sequence homology to Streptomyces sigma factors, has been known for a global regulatory protein stimulating multiple antibiotic biosynthetic pathways. Although the detailed regulatory mechanism of AfsK-AfsR-AfsR2 system has been well characterized, very little information about the AfsR2-dependent down-stream regulatory genes were characterized. Recently, the null mutant of afsS in S. coelicolor (the identical ortholog of afsR2) has been characterized through DNA microarray system, revealing that afsS deletion regulated several genes involved in antibiotic biosynthesis as well as phosphate-starvation. Through comparative DNA microarray analysis of afsR2-overexpressed S. lividans, here we also identify several afsR2-dependent genes involved in phosphate starvation, morphological differentiation, and antibiotic regulation in S. lividans, confirming that the AfsR2 plays an important pleiotrophic regulatory role in Streptomyces species.