• Fisheries and Aquatic Sciences
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• v.6 no.3
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• pp.116-124
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• 2003

Isolation and cloning of seven-band grouper (Epinephelus septemfasciatus) growth hormone cDNA from pituitary gland revealed an open reading frame of 612 bp coding for a pre-growth hormone of 204 amino acids with a 17 amino acid putative signal peptide. Deduced amino acid sequence showed that there was one possible N-glycosylation site at $Asn^{l84}$ and four cysteine residues $(Cys^{52},\;Cys^{160},\;Cys^{177},\;Cys^{185})$ on t e same positions as in some other species where they were involved in the stabilization of the tertiary structure. The seven-band grouper growth hormone (sbgGH) presented a $99.5\%$ amino acid sequence identity with the growth hormone of Epinephelus coioides and contained the conserved hormone domain region. Comparison of growth hormone sequences from evolutionarily diverse species revealed 25 amino acid residues conserved in jawless fishes to modern mammals. It also revealed an evolutionary trend to retain the same polypeptide sequence even in the distantly related animals while allowing alterations to occur in polypeptides of the closely related species. In order to create a recombinant system to produce high levels of the growth hormone, it was expressed in Escherichia coli (BL21) cells. The gel analysis revealed theoretically expected molecular weights for both mature and pre-sbgGHs.

• Journal of Microbiology and Biotechnology
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• v.12 no.5
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• pp.773-779
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• 2002

A gene (hap) encoding aminopeptidase from the chromosomal DNA of Bacillus licheniformis was cloned. The gene is 1,347 bp long and encodes a 449 amino acid preproprotein with a major mature region of 401 amino acids (calculated molecular mass 43,241 Da). N-Terminal sequence of the purified protein revealed a potential presence of N-terminal propeptide. The deduced primary amino acid sequence and the mass analysis of the purified protein suggested that a C-terminal peptide YSSVAQ was also cleaved off by a possible endogeneous protease. Tho amino acid sequence displayed 58% identity with that of the aminopeptidase from alkaliphilic Bacillus halodurans. This bacterial enzyme was overexpressed in recombinant Escherichia coli and Bacillus subtilis cells. Clones containing the intact hap gene, including its own promoter and signal sequence, gave rise to the synthesis of extracellular and thrmostable enzyme by B. subtilis transformants. The secreted protein exhibited the same biochemical properties and the similar apparent molecular mass as the B. lichenzyormis original enzyme.

• Fisheries and Aquatic Sciences
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• v.20 no.6
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• pp.10.1-10.7
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• 2017

Small GTPases are well known as one of the signal transduction factors of immune systems. The ADP-ribosylation factors (ARFs) can be classified into three groups based on the peptide sequence, protein molecular weight, gene structure, and phylogenetic analysis. ARF1 recruits coat proteins to the Golgi membranes when it is bound to GTP. The class I duplicated ARF gene was cloned and characterized from the olive flounder (Paralichthys olivaceus) for this study. PoARF1b contains the GTP-binding motif and the switch 1 and 2 regions. PoARF1b and PoARF1b mutants were transfected into a Hirame natural embryo cell to determine the distribution of its GDP/GTP-bound state; consequently, it was confirmed that PoARF1b associates with the Golgi body when it is in a GTP-binding form. The results of the qPCR-described PoARF1b were expressed for all of the P. olivaceus tissues. The authors plan to study the gene expression patterns of PoARF1b in terms of immunity challenges.

• Food Science of Animal Resources
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• v.33 no.3
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• pp.331-334
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• 2013

Osteopontin (OPN) is a secreted phosphorylated glycoprotein. It has an important role in multiple biological processes including cell survival, bone remodeling, inhibition of ectopic calcification, as well as, is thought to have potential immune modulation activities. In this work, we isolated and characterized a full-length open reading frame (ORF) of Korean native cow's OPN from Korean native cow's (Hanwoo) kidney, and successfully cloned firstly on Hanwoo's OPN. The sequencing results indicated that the isolated cDNA was 1190 bp in length containing a complete ORF of 837 bp. It encoded a precursor protein Hanwoo's OPN consisting of 278 amino acids with a signal peptide of 16 amino acids. Amino acid homology was found to be 99.3% as compared to the corresponding sequences of Holstein bone marrow OPN. Hanwoo's kidney OPN and Holstein bone marrow OPN are different only in two amino acid residues 42 and 56, amino acid residue 42 is Thr (T) ${\leftrightarrow}$ Ile (I), and amino acid residue 56 is Ala (A) ${\leftrightarrow}$ Thr (T) respectively. These results from the present work would be helpful to elucidate the biological function of Hanwoo's OPN and provided a foundation for further insight into role of Hanwoo's OPN.

• Applied Biological Chemistry
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• v.32 no.1
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• pp.64-72
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• 1989

Near full length cDNA clones encoding the rice seed storage protein, prolamine, were isolated and divided into two homology classes based on cross-hybridization and DNA sequencing analysis. These cDNA clones contain a single open reading frame encoding a putative rice prolamine precursor(M.W.=17,200) possessing atypical 14 amino acid signal peptide. Clones of these two homology classes diverge mainly by insertions/deletions of short nucleotide stretches and point mutations. The deduced primary structures of both types of prolamine polypeptides are devoid of any major tandem repetitive sequences, a feature prevalent in other cereal prolamines. No significant homology teas detected between the rice prolamine and other cereal prolamines, indicating that the rice gene evolved from a different ancestor that gave rise to other cereal prolamine genes. Developing wheat and rice endosperms were examined using ultrathin sections prepared from tissues harvested at various days after flowering. By immunocytochemical localization techniques, wheat prolamines are localized within vesicles from Golgi apparatus and in homogeneous regions of protein bodies. The involvement of the goli apparatus in the packaging of wheat prolamines into protein bodies indicates a pathway which differs from the mode of other cereal prolamines and resembles the mechanism employed for the storage of rice glutelin and legume globulins.

• Journal of Microbiology and Biotechnology
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• v.19 no.6
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• pp.610-615
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• 2009

As a provirus, polydnavirus has a segmented DNA genome on chromosome(s) of host wasp. It contains several genes in each segment that presumably play critical roles in regulating physiological processes of target insect parasitized by the wasp. A cysteine-rich protein 1 (CRP1) is present in the polydnavirus Cotesia plutellae bracovirus (CpBV) genome, but its expression and physiological function in Plutella xylostella parasitized by the viral host C. plutellae is not known. This CpBV-CRP1 encoding 189 amino acids with a putative signal peptide (20 residues) was persistently expressed in parasitized P. xylostella with gradual decrease at the late parasitization period. Expression of CpBV-CRP1 was tissue-specific in the fat body/epidermis and hemocyte, but not in the gut. Its physiological function was analyzed by inducing transient expression of a CpBV segment containing CpBV-CRP1 and its promoter, which caused significant reduction in hemocyte -spreading and delayed larval development. When the treated larvae were co-injected with double-stranded RNA of CpBV-CRP1, the expression of CpBV-CRP1 disappeared, whereas other genes encoded in the CpBV segment was expressed. These co-injected larvae significantly recovered the hemocyte-spreading capacity and larval development rate. This study reports that CpBV-CRP1 is expressed in P. xylostella parasitized by C. plutellae and its physiological function is to alter the host immune and developmental processes.

• Journal of Microbiology
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• v.37 no.4
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• pp.234-242
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• 1999

To obtain the recombinant circumsporozoite (CS) protein for the diagnosis of patients and seroepidemiology of Plasmodium vivax malaria which have been prevalent in northern part of Kyonggido, the CS protein gene was amplified by the polymerase chain reaction (PCR) from genomic DNA of the Korean vivax malaria patient. The gene consists of 1,123 nucleotides except signal peptide sequences and had an uninterrupted reading frame encoding a protein of 374 amino acids with a central region of 20 tandem repeats of the nonapeptide. The CS protein gene was expressed in Escherichia coli and purified, the molecular weight of recombinant CS protein was about 44 kDa (monomer) under denaturing purification and about 65 kDa (dimer) under native purification by SDS-PAGE. The purified recombinant CS protein which has antigenicity to malaria patients in Western blot analysis and Enzyme-linked immunosorbent assay, reacted only with the serum of P. vivax (PV210) infected malaria patients with no cross reaction to the P. falciparum malaria patient. The recombinant CS protein purified in this study will serve as a useful antigen to support the diagnosis of malaria patients and seroepidemiology.

• ALGAE
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• v.24 no.2
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• pp.85-91
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• 2009

The filamentous cyanobacterium Anabaena sp. Strain PCC 7120 produces a developmental patten of single hete- rocysts separated by approximately 10 vegetative cells. Heterocysts differentiate from vegetative cells and are spe- cialized for nitrogen fixation. The patS gene, which encodes a small peptide that inhibits heterocyst differentiation, is expressed in proheterocysts and plays a critical role in establishing the heterocyst pattem. Another key regulator of heterocyst development is the hetR gene. hetR mutants fail to produce heterocysts and extra copies of hetR on a plas- mid cause a multiple contiguous heterocyst phenotype. To elucidate the relationship between these two counter act- ing factors in the genetic regulatory pathway during heterocyst differentiation, the expression patterns of a patS-gfp and a hetR-gfp fusion were examined in a patS deletion and a hetR deletion strain. The results, in combination with the result from a hetR and patS double deletion strain, suggest patS and hetR are mutually antagonistic and the bal- ance between these two factors in tow different cell types (heterocysts and vegetative cells) may be critical during the decision making process on their cell fates.

• Journal of the Korean Magnetic Resonance Society
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• v.13 no.2
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• pp.108-116
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• 2009

The HP0902, a homodimeric 22.1 kDa protein, has been suggested as a putative secretory protein from Helicobacter pylori, although the protein possesses no signal peptide for secretion. Since it may be associated with the virulence of the bacterium, NMR study has been initiated in terms of structural genomics. In our previous effort to assign the backbone NMR resonances, using 800MHz NMR machine at pH 7.8, the resonances from eight of the 99 residues could not be assined due to missing of the signals. In this work, to enhance the extent of assignments, a 900 MHz machine was employed and the sample pH was reduced down to 6.5. Finally, almost all signals, except for those from G9 and S24, could be clearly assigned. The determined secondary structure using the assined chemical shifts indicated that the HP0902 consists of 11 ${\beta}$-strands with no helices. In our database search result, HP0902 was predicted to interact with VacA (Vacuolating cytotoxin A), which is a representative virulence factor secreted from Helicobacter pylori. Thus, molecular interaction between HP0902 and VacA would be worthy of investigation, on the basis of the present results of NMR assignments.

• Journal of agriculture & life science
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• v.46 no.2
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• pp.129-137
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• 2012

A carboxymethyl cellulase gene, cel5B, was cloned, sequenced, and expressed in Escherichia coli. pRCS20 in E. coli was identified from metagenomic cosmid library of cow rumen for cellulase activity on a carboxymethyl cellulose agar plates. Cosmid clone (RCS20) was partially digested with Sau3AI, ligated into BamHI site of pBluescript II SK+ vector, and transformed into E. coli $DH5{\alpha}$. The insert DNA of 1.3 kb was obtained, designated cel5B, which has the activity of hydrolyzation of CMC. The cel5B gene had an open reading frame (ORF) of 1,059 bp encoding 352 amino acids with a signal peptide of 48 amino acids and the conserved region, VIYEIYNEPL, belongs to the glycosyl hydrolase family 5. The molecular mass of Cel5B protein expressed from E. coli $DH5{\alpha}$ exhibited to be about 34 kDa by CMC-SDS-PAGE. The optimal pH was 8.0, and the optimal temperature was about $50^{\circ}C$ for its enzymatic activity.