This study aimed to develop a UPLC-MS/MS method for determining plasma levels of L-aspartic acid and L-asparagine and the activity of L-asparaginase. L-aspartic acid, L-asparagine, and L-aspartic acid-2,3,3-d3 were extracted from human plasma by protein precipitation with sulfosalicylic acid (30%, v/v). The plasma samples were analyzed using an Imtakt Intrada amino acid analysis column with 25 mM ammonium formate and 0.5% formic acid in acetonitrile as the mobile phase with step gradient method at a flow rate of 0.5 mL/min. The injection volume was 5 µL, and the total run time was 15 min. Inter- and intra-batch accuracies (%) ranged from 96.62–106.0% for L-aspartic acid and 89.85–104.8%, for L-asparagine, and the coefficient of variation (CV%) did not exceed 7%. The validation results for L-aspartic acid and L-asparagine satisfied the specified criterion, however, the results for L-asparaginase activity assay showed a borderline validity. This study could be a foundation for further development of therapeutic drug monitoring systems using UPLC-MS/MS.
Ammonium Compounds ; Asparagine ; Aspartic Acid ; Drug Monitoring ; Humans ; Methods ; Plasma

Human papillomavirus (HPV) 16, E7 proteins derived from the prototype (Bac73) and natural variant (Bac101) E7 open reading frame were produced in Sf9 insect cells. The variant E7 gene occurred naturally by substitution mutation at the position of 88 nucleotide, resulting serine instead of asparagine. Using E7 specific monoclonal antibody (VD6), both E7 proteins were identified in recombinant baculovirus infected SF9 cells. Radiolabelling and immunoprecipitation analysis revealed that both E7 proteins were phosphoproteins. Immunostaining result showed that E7 proteins were mainly localized in the cytoplasm. Nuclear form of E7 proteins was also detected after a sequential fractionation procedure for removing chromatin structure. Considering that the VD6 recognition site in E7 protein is located within 10 amino acid at the N-terminus, this region appears to be blocked by the nuclear component. Western blot analysis revealed that nuclear form was more abundant than cytoplasmic E7 proteins. Time course immunostaining showed that the primary location of E7 protein was the nucleus and exported to the cytoplasm as proteins were accumulated. These events occurred similarly in both Bac73 and Bac101 infected Sf9 cells, suggesting that these two proteins may have similar biological functions.
Asparagine ; Baculoviridae* ; Blotting, Western ; Chromatin ; Cytoplasm ; Humans* ; Immunoprecipitation ; Insects ; Open Reading Frames ; Phosphoproteins ; Serine ; Sf9 Cells

PURPOSE: In spite of an extensive vaccination program, parvoviral infections still pose a major threat to the health of dogs. MATERIALS AND METHODS: We isolated a novel canine parvovirus (CPV) strain from a dog with enteritis. Nucleotide and amino acid sequence analysis of the isolate showed that it is a novel type 2b CPV with asparagine at the 426th position and valine at the 555th position in VP2. To develop a vaccine against CPV infection, we passaged the isolate 4 times in A72 cells. RESULTS: The attenuated isolate conferred complete protection against lethal homologous CPV infection in dogs such that they did not develop any clinical symptoms, and their antibody titers against CPV were significantly high at 7-11 days post infection. CONCLUSION: These results suggest that the virus isolate obtained after passaging can be developed as a novel vaccine against paroviral infection.
Animals ; Asparagine ; Dogs ; Enteritis ; Parvovirus, Canine ; Sequence Analysis, Protein ; Sprains and Strains ; Vaccination ; Vaccines ; Valine ; Viruses

This study was carried out to obtain the basic data for artificial culture of veiled lady mushroom (Dictyophora spp). The optimal conditions for the mycelial growth were 25degrees C and pH 5.0 for all isolates except the optimal temperature of 30degrees C for D. echinovolvata ASI 32002 and Phallus rugulosus . The optimal medium for Dictyophora spp. was PBA (potato bamboo sawdust extract agar) medium. The strain ASI 32002, D. echinovolvata , grew faster than. D. indusiata ASI 32003 and Phallus rugulosus ASI 25007 on the medium. Carbon sources such as glucose, maltose and inuline were favorable for stimulating a mycelial growth of the two strains of ASI 32002 and ASI 32003. Asparagine and glutamine appeared to be favorable to the strain ASI 32002 and ASI 32003, where as alanine, one of nitrogen source also favorable to the strain ASI 32002. The optimum C/N ratio of the two isolates of ASI 32002 and ASI 32003 was about 25 : 1 when 2% glucose as carbon source was mixed with the basal medium. While, in the case of 4% as carbon source, the optimum C/N ratio was about 30 : 1.
Agaricales* ; Alanine ; Asparagine ; Carbon ; Cultural Characteristics* ; Glucose ; Glutamine ; Hydrogen-Ion Concentration ; Inulin ; Maltose ; Nitrogen

In an effort to develop novel mushroom-derived anti-obesity nutraceuticals, water and ethanol extracts containing the lipaseinhibitory compound from Phellinus linteus were prepared, and their nutritional components were determined. The optimal conditions for the extraction of P. linteus lipase inhibitor involved the treatment of the fruiting bodies with distilled water at 80degrees C for 72 hr and 80% ethanol at 100degrees C for 60 hr, respectively. The distilled water extract and ethanol extract contained 10.9% and 6.11% of crude protein, and 0.96% and 15.86% of crude fat, respectively. Additionally, the distilled water extract contained a large quantity of minerals, including 239.5 mg of K, 39.3 mg of Mg, and 39.3 mg of Na. The free amino acid content of the distilled water extracts was also higher than that of the ethanol extracts, and in particular, the distilled water extracts contained 5,139 mg of asparagine, 3,891 mg of tryptophan, 2,598 mg of alanine, and 2,066 mg of serine in 100 g of the distilled water extracts. 100 g of the distilled water and ethanol extracts were found to contain 12.31 g and 8.16 g of malic acid, respectively.
Alanine ; Asparagine ; Dietary Supplements ; Ethanol ; Fruit ; Lipase ; Malates ; Minerals ; Serine ; Tryptophan ; Water

Usnea longissima has a long history of use as a traditional medicine. Several bioactive compounds, primarily belonging to the polyketide family, have been isolated from U. longissima. However, the genes for the biosynthesis of these compounds are yet to be identified. In the present study, three different types of non-reducing polyketide synthases (UlPKS2, UlPKS4, and UlPKS6) were identified from a cultured lichen-forming fungus of U. longissima. Phylogenetic analysis of product template domains showed that UlPKS2 and UlPKS4 belong to group IV, which includes the non-reducing polyketide synthases with an methyltransferase (MeT) domain that are involved in methylorcinol-based compound synthesis; UlPKS6 was found to belong to group I, which includes the non-reducing polyketide synthases that synthesize single aromatic ring polyketides, such as orsellinic acid. Reverse transcriptase-PCR analysis demonstrated that UlPKS2 and UlPKS4 were upregulated by sucrose; UlPKS6 was downregulated by asparagine, glycine, and alanine.
Alanine ; Asparagine ; Fungi* ; Glycine ; Humans ; Medicine, Traditional ; Polyketide Synthases ; Polyketides ; Sucrose ; Usnea*

Amino-acid neurotransmitter system dysfunction plays a major role in the pathophysiology of depression. Several studies have demonstrated the potential of amino acids as a source of neuro-specific biomarkers could be used in future diagnosis of depression. Only partial amino acids such as glycine and asparagine were determined from certain parts of rats' brain included hippocampi and cerebral cortex in previous studies. However, according to systematic biology, amino acids in different area of brain are interacted and interrelated. Hence, the determination of 34 amino acids through entire rats' brain was conducted in this study in order to demonstrate more possibilities for biomarkers of depression by discovering other potential amino acids in more areas of rats' brain. As a result, 4 amino acids (L-aspartic acid, L-glutamine, taurine and gamma-amino-n-butyric acid) among 34 were typically identified as potentially primary biomarkers of depression by data statistics. Meanwhile, an antidepressant called Fluoxetine was employed to verify other potential amino acids which were not identified by data statistics. Eventually, we found L-alpha-amino-adipic acid could also become a new potentially secondary biomarker of depression after drug validation. In conclusion, we suggested that L-aspartic acid, L-glutamine, taurine, gamma-amino-n-butyric acid and L-alpha-amino-adipic acid might become potential biomarkers for future diagnosis of depression and development of antidepressant.
Amino Acids ; Animals ; Asparagine ; Aspartic Acid ; Biomarkers ; Biology ; Brain* ; Cerebral Cortex ; Depression ; Diagnosis ; Fluoxetine ; Glutamine ; Glycine ; Neurotransmitter Agents ; Rats* ; Taurine

Coxsackievirus B3 (CVB3) is a non-enveloped virus that has a single-stranded RNA genome. CVB3 induces myocarditis, and ultimately, dilated cardiomyopathy. A myocarditis variant of CVB3 (CVB3 H3) and its antibody-escape mutant (CVB3 10A1) were studied previously; H3 was found to induce myocarditis and 10A1 was found to be attenuated in infected mice. Although amino acid residue 165, located in a puff region of VP2, was found to be altered (i.e., the H3 asparagine was altered to aspartate in 10A1), the detailed mechanism of attenuation was not clearly elucidated. Here, DNA microarray technology was used to monitor changes in mRNA levels of infected mouse hearts after CVB3 H3 and 10A1 infection. This tool was used to elucidate the pathogenic mechanisms of viral infection by understanding virus-host interactions. We identified several genes, including protein tyrosine kinases, Ddr2 and Ptk2, as well as Clqb and Crry, involved in complement reactions, which may be involved in these viral processes. Thus, gene profiling can provide an opportunity to understand host immune responses to viral infection for gene therapy and may contribute to the identification of the target gene that is modified during treatment of viral myocarditis.
Animals ; Asparagine ; Aspartic Acid ; Cardiomyopathy, Dilated ; Complement System Proteins ; Genetic Therapy ; Genome ; Heart* ; Mice* ; Myocarditis ; Oligonucleotide Array Sequence Analysis ; Protein-Tyrosine Kinases ; RNA ; RNA, Messenger

Bovine coronavirus (BCoV) causes severe diarrhea in newborn calves, and is associated with winter dysentery in adult cattle and respiratory infections in calves and feedlot cattle. Although the Korean BCoV vaccine strain, BC94, was isolated in 1995, there has still been no report of a molecular characterization of the vaccine strain. To characterize the vaccine strain, relationships between BC94 and field strains were investigated, based on sequence analysis and cross-immunity. We determined the complete sequences of the HE, N, and S genes from BC94 and four NVRQS isolates (SUN5, A3, 0501, 0502). Due to its major role in antigenicity, the spike proteins of the BCoVs were analyzed. BC94 showed distinctive genetic divergence from field isolates collected from 2002 to 2005. BC94, SUN5, and A3 had no virulence-specific sequence and there was a single amino acid change, from asparagine to lysine at residue 175, in the polymorphic region. Strains 0501 and 0502 had virulence-specific sequences at all seven sites. Although the recently isolated Korean BCoVs and BC94 were genetically different, the cleavage site of spike genes at 763~768 (KRRSRR) and the antigenic domain II of the spike protein, amino acid position 528, were conserved in all NVRQS isolates. The antigenic relatedness of KCD9, representative of recent Korean BCoVs, was compared with the Korean vaccine strain BC94. KCD9 showed cross-reactivity against BC94 by virus neutralization (VN) test. These results suggest that BC94 is antigenically closely related to field isolates and is still effective as a vaccine strain.
Adult ; Animals ; Asparagine ; Cattle ; Coronavirus, Bovine ; Diarrhea ; Dysentery ; Humans ; Infant, Newborn ; Korea ; Lysine ; Proteins ; Respiratory Tract Infections ; Sequence Analysis ; Sprains and Strains ; Viruses

Bovine coronavirus (BCoV) causes severe diarrhea in newborn calves, and is associated with winter dysentery in adult cattle and respiratory infections in calves and feedlot cattle. Although the Korean BCoV vaccine strain, BC94, was isolated in 1995, there has still been no report of a molecular characterization of the vaccine strain. To characterize the vaccine strain, relationships between BC94 and field strains were investigated, based on sequence analysis and cross-immunity. We determined the complete sequences of the HE, N, and S genes from BC94 and four NVRQS isolates (SUN5, A3, 0501, 0502). Due to its major role in antigenicity, the spike proteins of the BCoVs were analyzed. BC94 showed distinctive genetic divergence from field isolates collected from 2002 to 2005. BC94, SUN5, and A3 had no virulence-specific sequence and there was a single amino acid change, from asparagine to lysine at residue 175, in the polymorphic region. Strains 0501 and 0502 had virulence-specific sequences at all seven sites. Although the recently isolated Korean BCoVs and BC94 were genetically different, the cleavage site of spike genes at 763~768 (KRRSRR) and the antigenic domain II of the spike protein, amino acid position 528, were conserved in all NVRQS isolates. The antigenic relatedness of KCD9, representative of recent Korean BCoVs, was compared with the Korean vaccine strain BC94. KCD9 showed cross-reactivity against BC94 by virus neutralization (VN) test. These results suggest that BC94 is antigenically closely related to field isolates and is still effective as a vaccine strain.
Adult ; Animals ; Asparagine ; Cattle ; Coronavirus, Bovine ; Diarrhea ; Dysentery ; Humans ; Infant, Newborn ; Korea ; Lysine ; Proteins ; Respiratory Tract Infections ; Sequence Analysis ; Sprains and Strains ; Viruses