• Proceedings of the Korean Society of Applied Pharmacology
• /
• 1995.04a
• /
• pp.81-81
• /
• 1995

It is well known that Src Homology 2(SH2) domain in many intracellular signal transduction proteins is very important. The domain has about 100 amino acid residues and bind phosphotyrosine-containing peptide with high affinity and specificity. Lck SH2 domain is a Src-like, lymphocyte-specific tyrosine kinase. An 11-residue phosphopeptide derived from the hamster polvoma middle-T antigen, EPQp YEEIPIYL, binds with an 1 nM dissociation constant to Lck SH2 domain. And it is known that the phosphotyrosine and isoleucine residues of the peptide are tightly bound by two well-defined pockets on Lck SH2 domain's surface. To investigate the conformational changes during complexation of SH2 domain with phosphopeptide we have performed the molecular dynamics simulation for Lck SH2 domain with peptide and peptide-free form at look in aqueous solution. More than 3000 water molecules were incorporated to solvate Lck SH2 domain and peptide. Periodic boundary condition has been applied in molecular dynamics simulation. Data analysis with the results of that simulation shows that the phosphopeptide makes primary interaction with the Lck SH2 domain at six central residues, The comparison of the complexed and uncomplexed SH2 domain structures in solution has revealed only relatively small change. But the hydrophilic and hydrophobic pockets in the protein surface show the conformational changes in spite of the small structural difference between the complex and peptide-free forms.

• Journal of Animal Science and Technology
• /
• v.48 no.5
• /
• pp.671-682
• /
• 2006

This experiment was conducted to investigate the effects of dietary Ca levels and some feed additives such as isoflavone and casein phosphopeptide (CPP) on eggshell quality and hatching egg production in aged egg-type breeder hens. A total of three hundred and sixty, 56-week-old Hy-Line Brown breeder hens were divided into six groups and fed experimental diets of two levels of Ca (3.3% or 3.6%) either with addition of 0.2% isoflavone, 0.5% CPP or devoid of all for 5 weeks. There were no significant differences in laying performances and settable egg production among the groups. Significant increases (P<0.05) in eggshell strength were observed with increasing dietary Ca and addition of isoflavone, but not with addition of CPP. Fertility and hatchability were not influenced by dietary Ca and addition of isoflavone or CPP. The treatment had few significant effects on tibial proximal compositions and breaking strength. The concentrations of Ca, P, estrogen and calcitonin in serum were not affected by the dietary treatments. These results indicated that relatively high level of dietary Ca in combination with isoflavone had a beneficial effect on improving eggshell quality in aged egg-type breeder hens. But hatching egg production was not affected by dietary isoflavone or CPP.

• Bulletin of the Korean Chemical Society
• /
• v.35 no.3
• /
• pp.845-850
• /
• 2014

The rapid development of shotgun proteomics is paving the way for extensive proteome profiling, while providing extensive information on various post translational modifications (PTMs) that occur to a proteome of interest. For example, the current phosphoproteomic methods can yield more than 10,000 phosphopeptides identified from a proteome sample. Despite these developments, it remains a challenging issue to pinpoint the true phosphorylation sites, especially when multiple sites are possible for phosphorylation in the peptides. We developed the Phospho-UMC filter, which is a simple method of localizing the site of phosphorylation using unique mass classes (UMCs) information to differentiate phosphopeptides with different phosphorylation sites and increase the confidence in phosphorylation site localization. The method was applied to large scale phosphopeptide profiling data and was demonstrated to be effective in the reducing ambiguity associated with the tandem mass spectrometric data analysis of phosphopeptides.

• Molecules and Cells
• /
• v.23 no.3
• /
• pp.340-348
• /
• 2007

Although considerable effort has been devoted in the mass spectrometric analysis of phosphorylated peptides, successful identification of multi-phosphorylated peptides in enzymatically digested protein samples still remains challenging. The ionization behavior of multi-phosphorylated peptides appears to be somewhat different from that of mono- or di-phosphorylated peptides. In this study, we demonstrate increased sensitivity of detection of multi-phosphorylated peptides of beta casein without using phosphopeptide enrichment techniques. Proteinase K digestion alone increased the detection limit of beta casein multi-phosphorylated peptides in the LC-MS analysis almost 500 fold, compared to conventional trypsin digestion (~50 pmol). In order to understand this effect, various factors affecting the ionization of phosphopeptides were investigated. Unlike ionizations of phosphopeptides with minor modifications, those of multi-phosphorylated peptides appeared to be subject to effects such as selectively suppressed ionization by more ionizable peptides and decreased ionization efficiency by multi-phosphorylation. The enhanced detection limit of multi-phosphorylated peptides resulting from proteinase K digestion was validated using a complex protein sample, namely a lysate of HEK 293 cells. Compared to trypsin digestion, the numbers of phosphopeptides identified and modification sites per peptide were noticeably increased by proteinase K digestion. Non-specific proteases such as proteinase K and elastase have been used in the past to increase detection of phosphorylation sites but the effectiveness of proteinase K digestion for multi-phosphorylated peptides has not been reported.

• Bulletin of the Korean Chemical Society
• /
• v.28 no.7
• /
• pp.1195-1198
• /
• 2007

• Journal of Dairy Science and Biotechnology
• /
• v.38 no.3
• /
• pp.161-167
• /
• 2020

Cheese pizzas fortified with casein phosphopeptide (CPP) and calcium were subjected to an in vitro digestion to assess whether CPP could prevent the precipitation of calcium. The total calcium content of the cheese pizzas was adjusted to 1,000 mg per pizza (~370 g) with the addition of calcium originating from eggshells. Two levels of trypsin-digested caseins (367 and 459 mg), with a CPP content of ~20%, were added to each pizza. The in vitro digested pizzas were then centrifuged and the supernatant was mixed with Na₂HPO₃ at 37℃ to estimate the possible soluble effect of CPP on calcium. After 24 h of reaction, the solution was centrifuged and the calcium content in the resultant supernatant was analyzed by inductively coupled plasma-atomic emission spectroscopy. One-way statistical analyses showed that CPP had a positive effect on the solubilization of calcium against phosphate (p<0.05). Cheese pizza supplemented with 459 mg of CPP powder was able to prevent precipitation of calcium by 98.8%, whereas no CPP-added cheese pizza solubilized 86.4% of the calcium. A sensory test was also carried out, revealing that panelists could not discern the bitter taste of the CPP added to the pizzas.

• Journal of Korean society of Dental Hygiene
• /
• v.10 no.3
• /
• pp.473-481
• /
• 2010

Objectives : To evaluate the effect of fluoride application on the color and microhardness of bleached enamel and compare it to that of casein phosphopeptide-amorphous calcium phosphate (CPP-ACP) application. Methods : Twenty freshly extracted human adult molar were each sectioned into halves, the specimens divided and treated according to five experimental groups: Group 1, treatment with 10% carbamide peroxide (CP) bleaching agent; Group 2, treatment with 10% CP followed by a 1.23% fluoride gel application; Group 3, treatment with 10% CP followed by a 2.23% sodium fluoride varnish application; Group 4, treatment with 10% CP followed by a 0.11% sodium fluoride gel application; Group 5, treatment with 10% CP followed by a CPP-ACP gel application. All groups were treated 6 h per day for 14 days then immersed in distilled water for 2 weeks. Changes in enamel color were evaluated on Baseline and Day 14. Microhardness were evaluated on Baseline, Days 7 and 14. Statistical analysis was performed using one-way ANOVA and post-hoc Tukey tests. Results : All the bleached enamel specimens revealed increased whiteness and overall color value. Group 1 showed the lowest microhardness values than that of Groups 2, 3, 4 and 5. In all groups, the hardness of tooth after bleaching showed a significant decrease in the microhardness as compared with the one prior to tooth bleaching. The specimens treated with remineralizing agents showed relatively less reduction in enamel microhardness than control group. Conclusions : The addition of fluoride and CPP-ACP did not impede the whitening effect. The use of remineralizing agents during bleaching treatment can significantly enhance the microhardness of bleached enamel.

• Korean Journal of Environmental Agriculture
• /
• v.25 no.4
• /
• pp.371-381
• /
• 2006

New proteomic techniques have been pioneered extensively in recent years, enabling the high-throughput and systematic analyses of cellular proteins in combination with bioinformatic tools. Furthermore, the development of such novel proteomic techniques facilitates the elucidation of the functions of proteins under stress or disease conditions, resulting in the discovery of biomarkers for responses to environmental stimuli. The ultimate objective of proteomics is targeted toward the entire proteome of life, subcellular localization biochemical activities, and the regulation thereof. Comprehensive analysis strategies of proteomics can be classified into three categories: (i) protein separation via 2-dimensional gel electrophoresis (2-DE) or liquid chromatography (LC), (ii) protein identification via either Edman sequencing or mass spectrometry (MS), and (iii) proteome quantitation. Currently, MS-based proteomics techniques have shifted from qualitative proteome analysis via 2-DE or 2D-LC coupled with off-line matrix assisted laser desorption ionization (MALDI) and on-line electrospray ionization (ESI) MS, respectively, toward quantitative proteome analysis. In vitro quantitative proteomic techniques include differential gel electrophoresis with fluorescence dyes. protein-labeling tagging with isotope-coded affinity tags, and peptide-labeling tagging with isobaric tags for relative and absolute quantitation. In addition, stable isotope-labeled amino acids can be in vivo labeled into live culture cells via metabolic incorporation. MS-based proteomics techniques extend to the detection of the phosphopeptide mapping of biologically crucial proteins, which ale associated with post-translational modification. These complementary proteomic techniques contribute to our current understanding of the manner in which life responds to differing environment.

• BMB Reports
• /
• v.34 no.6
• /
• pp.537-543
• /
• 2001

In the present study, an in vitro ELISA system to assess the interaction between Src homology (SH)2 domains and phosphotyrosine that contain peptides was established using purified GST-conjugated SH2 proteins and synthetic biotinylated phosphotyrosine that contain oligopeptides. The SH2 domains bound the relevant phosphopeptides that were immobilized in the streptavidin-coated microtiter plate in a highly specific and dose-dependent manner. The epidermal growth factor receptor (EGFR)-, T antigen (T Ag)-, and platelet-derived growth factor receptor (PDGFR)-derived phosphopeptides interacted with the growth factor receptor binding protein (Grb)2/SH2, Lck/SH2, and phosphatidyl inositol 3-kinase (PI3K) p85/SH2, respectively. No cross-reactions were observed. Competitive inhibition experiments showed that a short phosphopeptide of only four amino acids was long enough to determine the binding specificity. Optimal concentrations of the GST-SH2 fusion protein and phosphopeptide in this new ELISA system for screening the binding blockers were chosen at 2nM and 500nM, respectively. When two candidate compounds were tested in our ELISA system, they specifically inhibited the Lck/SH2 and/or p85/SH2 binding to the relevant phosphopeptides. Our results indicate that this ELISA system could be used as an easy screening method for the discovery of specific binding blockers of protein-protein interactions via SH2 domains.

• Journal of the korean academy of Pediatric Dentistry
• /
• v.41 no.3
• /
• pp.218-224
• /
• 2014

This study was designed to evaluate the effectiveness of 5% sodium fluoride-polyvinyl alcohol (NaF-PVA) tape influencing enamel remineralization by analysing enamel surface microhardness (SMH) and variation of ${\Delta}F$ of QLF. After enamel demineralizing of specimen, these 60 specimens with average KHN of microhardness ranging from 50 to 100 and with ${\Delta}F$ of QLF ranging from -15 to -25 were divided into four groups : group 1 (control group), group 2 (NaF-PVA), group 3 (fluoride varnish, FluoroDose$^{(R)}$ varnish), group 4 (Casein phosphopeptide-amorphous calcium phosphate, Tooth mousse plus$^{TM}$). These specimens were treated with materials and then immersed in artificial saliva. We measured remineralization rate each using surface microhardness (SMH) and Quantitative light-induced fluorescence digital (QLF-D). As a result, NaF-PVA tape is better than group 1, 4 and have comparable remineralization effect with group 3 (p < 0.05).