• Bulletin of the Korean Chemical Society
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• v.27 no.4
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• pp.529-534
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• 2006

Chemical modification of the S. cerevisiae farnesyl protein transferase (FPT) with CMC, phenylglyoxal and DEPC resulted in enzyme inactivation, depending upon the reagent concentration. The peptide substrate GST-PEP-I, a GST-fused undecapeptide mimicking the C-terminus of $p21^{Ki-ras}$, protected the enzyme against inactivation by CMC which is specific to either aspartate or glutamate, while the other substrate farnesyl pyrophosphate (FPP) showed protection against phenylglyoxal which is the specific modifier of arginine residues, dependent on the substrate concentrations. Neither of the two substrates protected the enzyme against histidine inactivation by DEPC. It is suggested that there is at least one aspartate or glutamate residue at the peptide substrate binding site, and that at least one arginine residue is located at the binding site of FPP. There also seems to be at least one histidine residue which is critical for enzymic activity and is exposed toward the bulk solution, excluded from the substrate binding sites.

• Asian-Australasian Journal of Animal Sciences
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• v.27 no.12
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• pp.1705-1711
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• 2014

The objective of this study was to evaluate the effect of ruminally protected amino acids (RPAAs) and ruminally protected fat (RPF) supplementation on ruminal fermentation characteristics (in vitro) and milk yield and milk composition (in vivo). Fourteen mid-lactating Holstein dairy cows (mean weight $653{\pm}62.59kg$) were divided into two groups according to mean milk yield and number of days of postpartum. The cows were then fed a basal diet during adaptation (2 wk) and experimental diets during the treatment period (6 wk). Dietary treatments were i) a basal diet (control) and ii) basal diet containing 50 g of RPAAs (lysine and methionine, 3:1 ratio) and 50 g of RPF. In rumen fermentation trail (in vitro), RPAAs and RPF supplementation had no influence on the ruminal pH, dry matter digestibility, total volatile fatty acid production and ammonia-N concentration. In feeding trial (in vivo), milk yield (p<0.001), 4% fat corrected milk (p<0.05), milk fat (p<0.05), milk protein (p<0.001), and milk urea nitrogen (p<0.05) were greater in cows fed RPAAs and RPF than the corresponding values in the control group. With an index against as 0%, the rates of decrease in milk yield and milk protein were lower in RPAAs and RPF treated diet than those of basal diet group (p<0.05). In conclusion, diet supplemented with RPAAs and RPF can improve milk yield and milk composition without negatively affecting ruminal functions in Holstein dairy cows at mid-lactating.

• Asian-Australasian Journal of Animal Sciences
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• v.19 no.8
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• pp.1164-1173
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• 2006

Cellulase from Aspergillus niger, (${\alpha}$-amylase from Bacillus sp. and protease from Bacillus globigii were used as enzyme sources in this study to examine how their respective substrates protect them in two kinds of simulated gastrointestinal tract digesting processes. Avian total digest tract simulation test showed that filter paper, Avicel and cellulose resulted in 7.7, 6.4 and 7.4 times more activity than of unprotected cellulose, respectively. Protease with addition of casein, gelatin or soybean protein showed no significant protection response. Starch protected amylase to be 2.5 times activity of the unprotected one. Monogastric animal total tract digestion simulation test showed that filter paper, Avicel and cellulose resulted in 5.9, 9.0 and 8.8 times activity of unprotected cellulase, respectively. Casein, gelatin and soybean protein resulted in 1.2, 1.3 and 2.0 times activity of unprotected protease, respectively. Starch did not protect amylase activity in monogastric animal total tract simulation. Protection of mixed enzymes by substrates in two animal total tract simulation tests showed that filter paper in combination with soybean protein resulted in 1.5 times activity of unprotected cellulose, but all substrates tested showed no significant protection effect to protease. Soybean protein and starch added at the same time protected the amylase activity to be two times of the unprotected one. Test of non-purified substrate protection in two animal total digest tract simulation showed that cellulase activity increased as BSA (bovine serum albumin) concentration increased, with the highest activity to be 1.3 times of unprotected enzyme. However, BSA showed no significant protection effect to protease. Amylase activity increased to 1.5 times as BSA added more than 1.5% (w/v). Cellulase activity increased to 1.5 times as soybean hull was added higher than 1.5%. Amylase had a significant protection response only when soybean hull added up to 2%. Protease activity was not protected by soybean hull to any significant extent.

• Animal Bioscience
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• v.35 no.10
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• pp.1556-1565
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• 2022

Objective: Tan lambs (n = 36, 3 mo old, 19.1±0.53 kg) were used to assess effects of dietary guanidinoacetic acid (GAA) and rumen-protected methionine (RPM) on growth performance, carcass traits, meat quality, and serum parameters. Methods: Lambs were randomly assigned to three treatment groups, with 6 pens per group and 2 lambs per pen. Dietary treatments were: basal diet alone (I); basal diet supplemented with 0.08% GAA+0.06% RPM (II); and basal diet supplemented with 0.08% GAA+0.08% RPM (III). Diets were provided three times a day for 90 d. Intake per pen was recorded daily and individual lamb body weight (BW) was measured monthly. Carcass traits were measured after slaughter and meat quality at the end of the experiment, blood samples were taken on a subgroup of lambs for analysis of indicators mostly related to protein metabolism. Results: Final BW and average daily gain for the first and second month, and for the entire experiment were greater in Treatment II compared to Treatment I (p<0.05), whereas feed to gain ratio was lower (p<0.05). Treatment II had the optimal dressing percentage and net meat weight proportion, as well as crude protein and intramuscular fat concentrations in muscles. Treatment II improved meat quality, as indicated by the greater water holding capacity, pH after 45 min and 48 h, and lower shear force and cooking loss. Dietary supplementation of GAA and RPM also increased the meat color a^* and b^* values at 24 h. Finally, Treatment II increased total protein, and serum concentrations of albumin and creatinine, but decreased serum urea nitrogen concentrations, indicating improved protein efficiency. Conclusion: In this study, 0.08% GAA+0.06% RPM supplementation improved growth performance and meat quality of Tan lambs.

• BMB Reports
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• v.29 no.5
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• pp.436-441
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• 1996

The palmitoyl acyl carrier protein (ACP) specific thioesterase (EC 3.1.2.14) from Iris pseudoacorus was purified and characterized. The thioesterase which was very unstable in relatively high salt concentrations was eluted using a co-gradient of Triton X-100 and low concentration of KCl or Na-phosphate from Q-Sepharose, DEAE-Sepharose, and hydroxyapatite chromatography. SDS-PAGE analysis showed a single band with a molecular weight of 35,000. The native molecular weight of approximately 37,000 was estimated by Sephacryl S-200 chromatography, indicating that the enzyme is a monomer. The thioesterase activity was inhibited about 75% and 50% by N-ethylmaleimide (2 mM) and phenylmethylsulfonyl fluoride (2 mM). respectively. The N-ethylmaleimide-inactivation was protected by sodium palmitate but the inactivation with phenylmethylsulfonyl fluoride was not protected. Oxidation of thiols by 2 mM 5.5'-dithio-bis-(2-nitrobenzoic acid) resulted in 65% inactivation of the enzyme. These results suggest that a cysteinyl residue is essential to the catalytic reaction of the enzyme. The enzyme activity was increased by sodium citrate and also by $Cu^{2+}$

• Asian-Australasian Journal of Animal Sciences
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• v.22 no.5
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• pp.659-666
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• 2009

Metabolizable protein (MP) supply and amino acid balance in the intestine were manipulated through selection of highly digestible rumen-undegradable protein (RUP) sources and protected methionine (Met) supplementation. Four ruminallycannulated, multiparous Holstein cows averaging 193${\pm}$13 days in milk were used in a 4${\times}$4 Latin square design to assess N utilization and milk production responses to changes in RUP level, post-ruminal RUP digestibility and protected Met supplementation. Treatments were A) 14.0% crude protein (CP), 8.0% rumen degradable protein (RDP) and 6.0% RUP of low intestinal digestibility (HiRUP-LoDRUP); B) 14.1% CP, 8.1% RDP and 6.0% RUP of high intestinal digestibility (HiRUP-HiDRUP); C) 13.1% CP, 7.9% RDP and 5.2% RUP of high intestinal digestibility (LoRUP-HiDRUP), and D) 13.1% CP, 7.9% RDP and 5.2% RUP of high intestinal digestibility plus rumen escape sources of Met (LoRUP-HiDRUP+Met). Experimental diets were formulated to have similar concentrations of RDP, net energy of lactation ($NE_L$), neutral detergent fiber (NDF), acid detergent fiber (ADF), calcium, phosphorus and ether extract using the NRC model (2001). Results showed that dry matter intake (DMI), production of milk fat and protein were similar among treatments. Milk production was similar for diet HiRUP-LoDRUP, HiRUP-HiDRUP and LoRUP-HiDRUP+Met, and significantly higher than diet LoRUP-HiDRUP. Milk fat and protein percentage were higher for cows receiving HiDRUP treatments, with the greatest increases in the diet LoRUP-HiDRUP+Met. There was no significant change in ruminal pH, $NH_3g-N$ and volatile fatty acid (VFA) concentration among all treatments. Apparent digestibility of dry matter (DM), CP, NDF and ADF and estimated bacterial CP synthesis were similar for all treatments. Nitrogen intakes, blood and milk urea-N concentrations were significantly higher for cows receiving HiRUP diets. Urine volume and total urinary N excretion were significantly lowered by LoRUP diets. Lowering dietary RUP level while supplementing the highly digestible RUP source with rumen escape sources of Met resulted in similar milk production, maximal milk fat and protein concentration and maximum N efficiency, indicating that post-ruminal digestibility of RUP and amino acid balance in the small intestine can be more important than total RUP supplementation.

• BMB Reports
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• v.46 no.10
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• pp.495-500
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• 2013

Turnip yellow mosaic virus (TYMV) is a positive strand RNA virus. We have modified TYMV coat protein (CP) by inserting a c-Myc epitope peptide at the N- or C-terminus of the CP, and have examined its effect on assembly. We introduced the recombinant CP constructs into Nicotiana benthamiana leaves by agroinfiltration. Examination of the leaf extracts by agarose gel electrophoresis and Western blot analysis showed that the CP modified at the N-terminus produced a band co-migrating with wild-type virions. With C-terminal modification, however, the detected bands moved faster than the wild-type virions. To further examine the effect, TYMV constructs producing the modified CPs were prepared. With N-terminal modification, viral RNAs were protected from RNase A. In contrast, the viral RNAs were not protected with C-terminal modification. Overall, the results suggest that virion assembly and RNA packaging occur properly when the N-terminus of CP is modified, but not when the C-terminus is modified.

• Archives of Plastic Surgery
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• v.40 no.5
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• pp.517-521
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• 2013

Background Diabetes is characterized by chronic hyperglycemia, which can increase reactive oxygen species (ROS) production by the mitochondrial electron transport chain. The formation of ROS induces oxidative stress and activates oxidative damage-inducing genes in cells. No research has been published on oxidative damage-related extracellular superoxide dismutase (EC-SOD) protein levels in human diabetic skin. We investigated the expression of EC-SOD in diabetic skin compared with normal skin tissue in vivo. Methods The expression of EC-SOD protein was evaluated by western blotting in 6 diabetic skin tissue samples and 6 normal skin samples. Immunohistochemical staining was also carried out to confirm the EC-SOD expression level in the 6 diabetic skin tissue samples. Results The western blotting showed significantly lower EC-SOD protein expression in the diabetic skin tissue than in the normal tissue. Immunohistochemical examination of EC-SOD protein expression supported the western blotting analysis. Conclusions Diabetic skin tissues express a relatively small amount of EC-SOD protein and may not be protected against oxidative stress. We believe that EC-SOD is related to the altered metabolic state in diabetic skin, which elevates ROS production.

• BMB Reports
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• v.35 no.6
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• pp.590-594
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• 2002

To elucidate the effect of gamma-irradiation on the molecular properties of myoglobin, the secondary and tertiary structures, as well as the molecular weight size of the protein, were examined after irradiation at various irradiation doses. Gamma-irradiation of myoglobin solutions caused the disruption of the ordered structure of the protein molecules, as well as degradation, cross-linking, and aggregation of the polypeptide chains. A SDS-PAGE study indicated that irradiation caused initial fragmentation of the proteins and subsequent aggregation, due to cross-linking of the protein molecules. The effect of irradiation on the protein was more significant at lower protein concentrations. Ascorbic acid protected against the degradation and aggregation of proteins by scavenging oxygen radicals that are produced by irradiation. A circular dichroism study showed that an increase of the irradiation decreased the a-helical content of myoglobin with a concurrent increase of the aperiodic structure content. Fluorescence spectroscopy indicated that irradiation increased the emission intensity that was excited at 280 nm.

• BMB Reports
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• v.37 no.2
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• pp.260-267
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• 2004

Transducin (T), the heterotrimeric guanine nucleotide binding protein in rod outer segments, serves as an intermediary between the receptor protein, rhodopsin, and the effector protein, cGMP phosphodiesterase. Labeling of T with dansyl chloride (DnsCl) inhibited its light-dependent guanine nucleotide binding activity. Conversely, DnsCl had no effect on the functionality of rhodopsin. Approximately 2-3 mol of DnsCl were incorporated per mole of T. Since fluoroaluminate was capable of activating DnsCl-modified T, this lysine-specific labeling compound did not affect the guanine nucleotide-binding pocket of T. However, the labeling of T with DnsCl hindered its binding to photoexcited rhodopsin, as shown by sedimentation experiments. Additionally, rhodopsin completely protected against the DnsCl inactivation of T. These results demonstrated the existence of functional lysines on T that are located in the proximity of the interaction site with the photoreceptor protein.