• Nutritional Sciences
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• v.5 no.2
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• pp.53-59
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• 2002

This study was conducted to examine the effects of dietary sources of fatty acids and protein on serum protein profiles, hepatic functional enzyme activities, mammary tumor incidence and tumor weight in 7, 12-dimethylbenz($\alpha$)anthracene (DMBA)-treated rats. The sources of dietary fatty acids were 18n6 (rich in linoleic acid), 18n3 (rich in linolenic acid) and 22n3 (rich in DHA) : sources of dietary protein were casein (C) and soy protein isolate (S). mammary tumors (MTs) were chemically induced by DMBA (9 mg/100 g body weight) which was gastrically intubated at 7 weeks of age. Each experimental diet was given for the following 25 weeks. Casein-fed rats (group C) exhibited significantly higher levels of weight gain and FER (food efficiency ratio) than did group S. Group C showed higher levels of serum protein and globulin, and higher albumin/globulin (A/G) ratios than group S. Liver functional enzyme activities (GOT, GPT, ALP, LDH, $\gamma$-GT) and LDH/GOT ratios were not influenced by dietary protein. GPT activity was lower in the group given 18n3, and ALP activity was lower in the group given 18n6. The incidence and total number of MTs appeared to be lower in the group given 22n3 than in the group given 18n3 or 18n6, even though the average weight of MTs was highest in the group given 22n3, The average weight of MTs was higher in the C group than in the S group. MT incidence had a positive correlation with LDH activity and LDH/GOT ratio. The average weight of MTs had a negative correlation with serum albumin levels and A/G ratios, and a positive correlation with ALP activity. This research suggests that the measurement of serum protein profiles and liver functional enzyme activities may be utilized to monitor the development of mammary tumors.

• Proceedings of the Korean Society for Applied Microbiology Conference
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• 1986.12a
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• pp.505.1-505
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• 1986

The gene for iso-1-cytochrome c for Saccharomyces cerevisiae was recloned into a pSP65 vector containing an active bacteriophage SP6 promoter. The iso-1-cytochrome c gene was cloned as an 856 bp Xho 1-Hind III fragment. When the resulting plasmid was digested at the Hind 111 site 279 bases downstream from the termination codon of the gene and transcribed in vitro using SP6 RNA polymerase, full length transcripts were produced. The SP6 iso-1-cytochrome c mRNA was translated using a rabbit reticulocyte lysate system and the protein products analyzed on SDS polyacrylamide gels. One major band was detected by autofluorography. This band was found to have a molecular weight of 12,000 Da and coincided with the Coomassie staining band of apocytochrome c from S. cerebisiae. The product was also shown to be identical with that of standard yeast apocytochrome c on an isoelectric focusing gel. The in vitro synthesized iso-a-cytochrome c was methylated by adding partially purified S-adenosyl-L-methionine . protein-lysine N-methyltransferase (Protein methylase III; EC 2.1.1.43) from S. cerevisiae along with S-adenosyl-L-methionine to the in vitro translation mixtures. The methylation was shown to be inhibited by the addition of the methylase inhibitor S-adenosyl-L-homocysteine or the protein synthesis inhibitor pu omycin. The methyl derivatives in the protein were identified as $\varepsilon$-N-mono, di and trimethyllysine by amino acid analysis. The molar ratio of methyl groups incorporated to that of cytochrome c molecules synthesized showed that 23% of the translated cytochrome c molecules were methylated by protein methylase III.

• Biomedical Science Letters
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• v.2 no.2
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• pp.231-240
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• 1996

Human blood clotting (coagulation) factor 9 cDNA which codes for 461 amino acid has been cloned by screening human fetal liver cDNA library using PCR. This 1.4 kb cDNA spanning from the ATG initiation codon to the TAA termination codon was cloned into bacterial .expression vector pGEX-2T, generating pGEX-F9 plasmid. The plasmid pGEX-F9 expresses about 73 kDa GST (Glutathione S-transferase)-Factor 9 fusion protein when introduced into E. coli. Western blot analysis using polyclonal antibody raised against human factor 9 confirmed this fusion protein contains factor 9 protein. The level of GST-factor 9 expression was about 20% of total protein and the purification of fusion protein was efficiently achieved by using GST agarose bead based on one step purification protocol.

• Asian-Australasian Journal of Animal Sciences
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• v.17 no.9
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• pp.1296-1302
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• 2004

This study was conducted to evaluate the possible application of 2 D-SDS-PAGE (2 DE)-based proteome analysis techniques to the assessment of extreme proteolysis in postmortem skeletal muscle. Eight Hanwoo longissimus muscles were incubated immediately after slaughter for 24 h at 5$^{\circ}C$, 15$^{\circ}C$ or 36$^{\circ}C$. Warner Bratzler (WB)-shear force and ultrastructural configuration were determined at 24 h, and rate of proteolysis to 24 h was determined by 1 D-SDS-PAGE (1 DE) and 2 DE. In addition, tentative protein identification was performed from peptide mass fingerprints of MALDI-ToF analysis of major protein groups on 2 DE profiles. The result showed that although ultrastructural configuration was similar between the 5$^{\circ}C$ and 36$^{\circ}C$ treatments, meat at 5$^{\circ}C$ had higher WBshear force (approximately 5 kg greater). A higher rate of protein degradation at 36$^{\circ}C$ was observed based on Troponin-T degradation, 1 DE, and 2 DE analysis. This indicates that proteolysis during the early postmortem period was a significant determinant of shear force at 24 h. Little difference in proteolysis between 5$^{\circ}C$ and 15$^{\circ}C$ treatments was found based on classic 1 DE profile assessment. Meanwhile, considerable differences in the 2 DE profiles between the two treatments were revealed, with substantially higher rate of proteolysis at 15$^{\circ}C$ compared to 5$^{\circ}C$. Nuclease treatment improved 2 DE profile resolution. 400 ${\mu}$g and 600 ${\mu}$g of sample loading appeared to be appropriate for 24 cm pH 3-10 and pH 5-7 IPG strips, respectively. Protein detection and quantification of the 5$^{\circ}C$, 15$^{\circ}C$ and 36$^{\circ}C$ 2 DE profiles revealed 78, 163 and 232 protein spots respectively that were differentially modified in terms of their electrophoretic properties between approximately pI 5.3-7.7 with the molecular weight range of approximately 71-12 kDa. The current results demonstrated that 2 DE was a superior tool to 1 DE for characterising proteolysis in postmortem skeletal muscle.

• Journal of Microbiology and Biotechnology
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• v.18 no.5
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• pp.845-851
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• 2008

TolC is an outer membrane porin protein and an essential component of drug efflux and type-I secretion systems in Gram-negative bacteria. TolC comprises a periplasmic $\alpha$-helical barrel domain and a membrane-embedded $\beta$-barrel domain. TdeA, a functional and structural homolog of TolC, is required for toxin and drug export in the pathogenic oral bacterium Actinobacillus actinomycetemcomitans. Here, we report the expression of the periplasmic domain of TdeA as a soluble protein by substitution of the membrane-embedded domain with short linkers, which enabled us to purify the protein in the absence of detergent. We confirmed the structural integrity of the TdeA periplasmic domain by size-exclusion chromatography, circular dichroism spectroscopy, and electron microscopy, which together showed that the periplasmic domain of the TolC protein family fold correctly on its own. We further demonstrated that the periplasmic domain of TdeA interacts with peptidoglycans of the bacterial cell wall, which supports the idea that completely folded TolC family proteins traverse the peptidoglycan layer to interact with inner membrane transporters.

• Journal of the East Asian Society of Dietary Life
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• v.9 no.4
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• pp.435-441
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• 1999

This study was designed to investigate the effects of roasting temperatures on the antinutritional factors and functional properties of sesame protein isolates. The protein contents a sesame protein isolates (SPI) prepared at roasting temperatures of 5$0^{\circ}C$, 10$0^{\circ}C$, 15$0^{\circ}C$ and 20$0^{\circ}C$ were 65.5%, 66.6%. 68.9% and 64.1%, respectively. Total phenolic compounds, condensed tannin and phytate contents of SPI was increased from 5$0^{\circ}C$ to 20$0^{\circ}C$. From color measurements, higher roasting temperature decreased 'L' and 'b' values significantly, but 'a' value was increased. The bulk density, fat absorption and water absorption of SPI was increased as the temperature of roasting was increased. Sesame protein isolates prepared by roasting at 5$0^{\circ}C$, 10$0^{\circ}C$ and 15$0^{\circ}C$ had higher emulsifying activity than those prepared by roasting at 20$0^{\circ}C$. Foaming capacity of SPI was not changed by roasting up to 10$0^{\circ}C$, but this property was reduced dramatically when roasted at 20$0^{\circ}C$.

• Asian-Australasian Journal of Animal Sciences
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• v.12 no.8
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• pp.1205-1214
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• 1999

Whole lupinus albus seeds were pressure toasted at temperatures of 100, 118 and $136^{\circ}C$ for 3, 7, 15 and 30 min to study rumen degradation and post-rumen digestion and to determine optimal heating conditions for the Dutch dairy feed industry. In sacco nylon bag and mobile bag techniques were employed for rumen and intestine incubations to determine ruminal degradation characteristics and intestinal digestion of crude protein (CP) in 4 lactation rumen cannulated and 4 lactating intestinal cannulated Dutch dairy cows fed 47% hay and 53% concentrate according to Dutch dairy requirements. Measured rumen degradation characteristics were soluble fraction (S), undegradable fraction (U), potentially degradable fraction (D), lag time (T0) and rate of degradation (Kd) of insoluble but degradable fraction. Percentage bypass feed protein (BCP), ruminal microbial protein synthesized based on available nitrogen (N_MP) and that based on available energy (E_MP), true protein supplied to the small intestine (TPSI), truly absorbed BCP (ABCP), absorbed microbial protein (AVP) in the small intestine, endogenous protein losses in the digestion (ENDP), true digested protein in the small intestine (TAP or DVE in Dutch) and degraded protein balance (PDB or OEB in Dutch) were totally evaluated using the new Dutch DVE/OEB System. Pressure toasting decreased (p<0.001) rumen degradability of CP. It reduced S (p<0.05) and Kd (p=0.06), increased D (p<0.05) and U (p<0.01) but did not alter T0 (p>0.05), thus resulting in dramatically increased BCP (p<0.001) with increasing time and temperature from 73.7 (raw) up to 182.5 g/kg DM ($136^{\circ}C/15min$). Although rumen microbial protein synthesized based on available energy (E_MP) was reduced, true protein (microbial and bypass feed protein) supplied to the small intestine (TPSI) was increased (p<0.001) from 153.1 (raw) to 247.6 g/kg DM ($136^{\circ}C/15min$). Due to digestibility of BCP in the intestine not changing (p>0.05) average 87.8%, the absorbed BCP increased (p<0.001) from 62.3 (raw) to 153.7 g/kg DM ($136^{\circ}C/15min$). Therefore DVE value of true digested protein in the small intestine was significantly increased (p<0.001) from 118.9 (raw) to 197.0 g/kg DM ($136^{\circ}C/15min$) and OEB value of degraded protein balance was significantly reduced (p<0.001) from 147.2 (raw) to 63.1 g/kg DM ($136^{\circ}C/15min$). It was concluded that pressure toasting was effective in shifting degradation of CP of lupinus albus from the rumen to small intestine without changing intestinal digestion. Further studies are required on the degradation and digestion of individual amino acids and on the damaging effects of processing on amino acids, especially the first limiting amino acids.

• Journal of Nutrition and Health
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• v.27 no.5
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• pp.419-428
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• 1994

To investigate the effect of dietary menhaden oil on protein kinase C (PKC) activity and membrane phospholipid composition in epidermal cells, female BALB/C mice were fed either menhaden oil or corn oil with two different levels(5% or 20%) for 6 weeks. Membrane phosphatidycholine(PC) was decreased in menhaden oil-fed group. Eicosapentaenoic acid(EPA) and Docosahexaenoic acid(DHA) were only presented in the acyl chain of membrane phospholipid of menhaden oil-fed mice, so that membrane fluidity of the group could be different from the other group. Both cytosolic and membrane-associated PKC activity in epidermal cells were decreased in menhaden oil-fed mice when compare with corn oil-fed mice. Furthermore, rate of PKC transfer from cytosol to membrane in menhaden oil-fed group was not as fast as in corn oil-fed group. Based on these observations, dietary menhaden oil might act differently from other dietary fat in carcinogenesis.

• Journal of the Korean Magnetic Resonance Society
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• v.2 no.2
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• pp.131-140
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• 1998

The calcium-induced structural changes in the skeletal muscle regulatory protein troponin C (NTnC) involve a transition from a ‘closed’to an ‘open’structure with the concomitant exposure of a large hydrophobic interaction site for target proteins. Structural studies have served to define this conformational change and elucidate the mechanism of the linkage between calcium binding and the induced structural changes. There are now several structures of NTnC available from both NMR and X-ray crystallography. Comparison of the calcium bound structures reveals differences in the level of opening. We have considered the concept of a flexible open state of NTnC as a possible explanation for this apparent discrepancy. We also present simulations of the closed-to-open transition which are in agreement with the flexibility concept and with experimental energetics data.

• Journal of Integrative Natural Science
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• v.4 no.3
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• pp.222-226
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• 2011

Cyclic AMP receptor protein(CRP) is involved in the activation of many genes corresponding to catabolite enzymes in Escherichia coli. In this study, mutant CRP(S128A) was used to elucidate the effect of Ser 128 on the cAMP-induced structural change. Based on the protease digestion and thermal analysis, serine 128 in CRP affects the cAMP binding capability and then structural change of CRP protein. In addition, CD spectra in near UV region revealed that S128A CRP retained the sensitive conformation to thermal effect relative to that of wild-type CRP, in spite of identical Tm values in the absence of cAMP.