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1.
Article in Chinese | WPRIM | ID: wpr-878883

ABSTRACT

Nano-LC MS/MS was used to analyze trypsin digested deer-hide gelatin(DHG) samples, hydroxylation and O-glycosylation on lysine sites of DHG were comprehensive identified by using PEAKS Studio software. The sites, sorts and amounts of hydroxylation and O-glycosylation on Type Ⅰ collagen α1 chain(COL1 A1) and α2 chain(COL1 A2) of DHG were revealed. As a result, 5 284 peptides were identified from DHG samples, which were mainly from COL1 A1 and COL1 A2. Among these peptides, there were 449 peptides with hydroxylysine, 442 with galactosyl-hydroxylysine, 449 with glucosyl-galactosyl-hydroxylysine. The major modified sites of hydroxylation and O-glycosylation in DHG were shown as follow: α1-9 N and α2-5 N in N-telopeptides, α1-87, α1-174, α1-930, α2-87, α2-174, α2-933 in triple helix domain, and α1-16 C in C-telopeptides. These hydroxylation and O-glycosylation were correlated with the formation and stability of collagen molecules and collagen fibrils. It is feasible for the collagens and peptides dissolving from deer skin collagen fibrils under high temperature and pressure decocting, high temperature and pressure also might destroy inter-molecular covalent cross-linking and help those glycol-peptides formations. The present study provided ideas and strategies for the in-depth investigation on DHG chemical constituents, and showed good theoretical significance and application value.


Subject(s)
Animals , Deer/metabolism , Gelatin , Glycosylation , Hydroxylation , Lysine/metabolism , Protein Processing, Post-Translational , Tandem Mass Spectrometry
2.
Article in Chinese | WPRIM | ID: wpr-921528

ABSTRACT

Objective To obtain the proteome and acetylome profiles of livers in mice during normal aging.Methods We applied tandem mass tag labeling and liquid chromatography tandem mass spectrometry and achieved proteome and acetylome data in C57BL/6J male mice aged 2 and 18 months under physiological conditions.Results A total of 4712 proteins were quantified by proteome profiling,and 4818 acetylated sites in 1367 proteins by acetylome profiling.The proteome and acetylome revealed moderate differences in the livers of young and old mice.There were 195 differentially expressed proteins in the proteome and 113 differentially expressed acetylated sites corresponding to 76 proteins in the acetylome.Functional enrichment analysis for the proteome showed that aging-associated upregulated proteins were mainly involved in fatty acid metabolism,epoxygenase P450 pathway,drug catabolic process,organic hydroxy compound metabolic process,and arachidonic acid metabolic process,while the downregulated proteins were related to regulation of gene silencing,nucleosome assembly,protein heterotetramerization,response to interferon,protein-DNA complex assembly and other processes.For the acetylome,the proteins with aging-associated upregulated acetylated sites mainly participated in cofactor metabolism,small molecule catabolic process,ribose phosphate metabolic process,ribonucleotide metabolic process,and purine-containing compound metabolic process,while the proteins with downregulated acetylated sites were associated with sulfur compound metabolic process,response to unfolded protein,and amino acid metabolic process.Conclusion We profiled the proteome and acetylome of livers in mice during normal aging and generated datasets for further research on aging.


Subject(s)
Acetylation , Aging , Animals , Liver , Lysine/metabolism , Male , Mice , Mice, Inbred C57BL , Proteome/metabolism
3.
Article in English | WPRIM | ID: wpr-880485

ABSTRACT

Ubiquitination, an essential post-transcriptional modification (PTM), plays a vital role in nearly every biological process, including development and growth. Despite its functions in plant reproductive development, its targets in rice panicles remain unclear. In this study, we used proteome-wide profiling of lysine ubiquitination in rice (O. sativa ssp. indica) young panicles. We created the largest ubiquitinome dataset in rice to date, identifying 1638 lysine ubiquitination sites on 916 unique proteins. We detected three conserved ubiquitination motifs, noting that acidic glutamic acid (E) and aspartic acid (D) were most frequently present around ubiquitinated lysine. Enrichment analysis of Gene Ontology (GO) annotations and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathways of these ubiquitinated proteins revealed that ubiquitination plays an important role in fundamental cellular processes in rice young panicles. Interestingly, enrichment analysis of protein domains indicated that ubiquitination was enriched on a variety of receptor-like kinases and cytoplasmic tyrosine and serine-threonine kinases. Furthermore, we analyzed the crosstalk between ubiquitination, acetylation, and succinylation, and constructed a potential protein interaction network within our rice ubiquitinome. Moreover, we identified ubiquitinated proteins related to pollen and grain development, indicating that ubiquitination may play a critical role in the physiological functions in young panicles. Taken together, we reported the most comprehensive lysine ubiquitinome in rice so far, and used it to reveal the functional role of lysine ubiquitination in rice young panicles.


Subject(s)
Acetylation , Lysine/metabolism , Oryza/metabolism , Plant Proteins/metabolism , Protein Interaction Maps , Protein Processing, Post-Translational , Proteome/metabolism , Ubiquitin/metabolism , Ubiquitination
4.
Article in English | WPRIM | ID: wpr-880483

ABSTRACT

Protein lysine methylation is a prevalent post-translational modification (PTM) and plays critical roles in all domains of life. However, its extent and function in photosynthetic organisms are still largely unknown. Cyanobacteria are a large group of prokaryotes that carry out oxygenic photosynthesis and are applied extensively in studies of photosynthetic mechanisms and environmental adaptation. Here we integrated propionylation of monomethylated proteins, enrichment of the modified peptides, and mass spectrometry (MS) analysis to identify monomethylated proteins in Synechocystis sp. PCC 6803 (Synechocystis). Overall, we identified 376 monomethylation sites in 270 proteins, with numerous monomethylated proteins participating in photosynthesis and carbon metabolism. We subsequently demonstrated that CpcM, a previously identified asparagine methyltransferase in Synechocystis, could catalyze lysine monomethylation of the potential aspartate aminotransferase Sll0480 both in vivo and in vitro and regulate the enzyme activity of Sll0480. The loss of CpcM led to decreases in the maximum quantum yield in primary photosystem II (PSII) and the efficiency of energy transfer during the photosynthetic reaction in Synechocystis. We report the first lysine monomethylome in a photosynthetic organism and present a critical database for functional analyses of monomethylation in cyanobacteria. The large number of monomethylated proteins and the identification of CpcM as the lysine methyltransferase in cyanobacteria suggest that reversible methylation may influence the metabolic process and photosynthesis in both cyanobacteria and plants.


Subject(s)
Bacterial Proteins/metabolism , Lysine/metabolism , Methyltransferases/metabolism , Photosynthesis , Protein Processing, Post-Translational , Synechocystis/growth & development
5.
Ciênc. saúde coletiva ; 20(1): 75-84, 01/2015. graf
Article in English | LILACS | ID: lil-733155

ABSTRACT

This study sought to verify the records on file and the number of cases of attempted suicide among children and adolescents who were attended by Emergency Care health professionals in the municipality of Matozinhos, Minas Gerais, Brazil. Documentary and descriptive research was conducted, the data for which was collected by means of an investigation of Outpatient Records from 2008 to 2010. Of the 73,000 files evaluated, those dealing with cases of attempted suicide among children and adolescents between the age of 3 and 18 years were selected. It was revealed that the health professionals, particularly physicians and nurses, fail to register the cases appropriately, invalidating information about the problem and potential prevention measures. The conclusion reached was that underreporting and the discrepancy of the diagnoses which were not duly referred to the competent agencies require rethinking and reviewing medical practices, and taking a systematic and careful look to address the individual as a complex whole.


Neste estudo procurou-se verificar o registro e o número de casos de tentativa de suicídio entre crianças e adolescentes do município de Matozinhos, Minas Gerais, Brasil, que foram atendidos pelos profissionais de saúde do Pronto-Atendimento. Trata-se de uma pesquisa documental e descritiva, cuja coleta dos dados ocorreu por meio de investigação nas Fichas Ambulatoriais, no período de 2008 a 2010. Das 73.000 fichas levantadas, selecionaram-se aquelas que tratavam de casos de tentativa de suicídio entre crianças e adolescentes do município, com idades entre três e 18 anos. Percebeu-se que os profissionais de saúde, mais especificamente os médicos e enfermeiros, não registram os casos de forma adequada, inviabilizando a informação sobre o problema e as medidas de prevenção. Concluiu-se que a subnotificação, a discrepância dos diagnósticos e o não encaminhamento aos órgãos competentes exigem repensar e rever a prática médica e dirigir um olhar sistematizado e cuidadoso para perceber o sujeito como um todo complexo.


Subject(s)
Aldehydes/chemistry , Cytochromes c/chemistry , Mitochondrial Membranes/metabolism , Oxidative Stress/drug effects , Amino Acid Sequence , Cardiolipins/chemistry , Cardiolipins/metabolism , Cytochromes c/metabolism , Electron Transport Complex IV/metabolism , Hydrogen-Ion Concentration , Histidine/chemistry , Histidine/metabolism , Lysine/chemistry , Lysine/metabolism , Molecular Sequence Data , Molecular Weight , Oxidative Stress/physiology , Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization , Time Factors
6.
Indian J Biochem Biophys ; 2014 Dec ; 51 (6): 559-566
Article in English | IMSEAR | ID: sea-156537

ABSTRACT

In addition to well-known process of proteasome-mediated degradation of polyubiquitinated proteins, monoubiquitination of proteins is also an important post-translational modification that regulates various non-degradative cellular processes like protein trafficking, cellular signalling, DNA replication and DNA repair. We have previously characterized a multi-domain cycling sequence binding protein LdCSBP from Leishmania donovani, which binds specifically to a conserved CAUAGAAG octamer containing RNAs via its uniquely arranged CCCH type Zn-fingers and degrades them using its Smr endonuclease domain, indicative of its potential role in the turnover of the S-phase mRNAs. Remarkably, its riboendonuclease activity is inhibited due to the incorporation of a monoubiquitin residue in the ZnF domain, though the target Lys residue remains unknown. Here, we report through systematic mutation of Lys residue to Ala that Lys-413 in LdCSBP is the site of monoubiquitination. However, the amino acid motif around the target Lys in LdCSBP is not consensus with any previously known monoubiquitination site, though partial homology is observed with a subset of recently identified mammalian ubiquitination target sites. Interestingly, Lys-413 of LdCSBP is conserved in the homologous annotated proteins from the related kinetoplastida parasites, suggesting similar monoubiquitination-mediated regulation of RNA endonuclease activity in the organisms.


Subject(s)
Amino Acid Sequence , Binding Sites , Endonucleases/chemistry , Endonucleases/genetics , Endonucleases/metabolism , Leishmania donovani/cytology , Leishmania donovani/physiology , Lysine/chemistry , Lysine/genetics , Lysine/metabolism , Molecular Sequence Data , Protein Binding , Protein Interaction Domains and Motifs , Protozoan Proteins/metabolism , RNA-Binding Proteins , S Phase/physiology , Structure-Activity Relationship , Ubiquitination , Zinc Fingers
7.
Braz. j. med. biol. res ; 47(12): 1057-1061, 12/2014. graf
Article in English | LILACS | ID: lil-727658

ABSTRACT

Endogenous carbon monoxide (CO), which is produced by the enzyme heme oxygenase (HO), participates as a neuromodulator in physiological processes such as thermoregulation and nociception by stimulating the formation of 3′,5′-cyclic guanosine monophosphate (cGMP). In particular, the acute physical restraint-induced fever of rats can be blocked by inhibiting the enzyme HO. A previous study reported that the HO-CO-cGMP pathway plays a key phasic antinociceptive role in modulating noninflammatory acute pain. Thus, this study evaluated the involvement of the HO-CO-cGMP pathway in antinociception induced by acute stress in male Wistar rats (250-300 g; n=8/group) using the analgesia index (AI) in the tail flick test. The results showed that antinociception induced by acute stress was not dependent on the HO-CO-cGMP pathway, as neither treatment with the HO inhibitor ZnDBPG nor heme-lysinate altered the AI. However, antinociception was dependent on cGMP activity because pretreatment with the guanylate cyclase inhibitor 1H-[1,2,4] oxadiazolo [4,3-a] quinoxaline-1-one (ODQ) blocked the increase in the AI induced by acute stress.


Subject(s)
Animals , Male , Acute Pain/prevention & control , Carbon Monoxide/metabolism , Cyclic GMP/metabolism , Heme Oxygenase (Decyclizing)/metabolism , Nociceptive Pain/prevention & control , Stress Disorders, Traumatic, Acute/metabolism , Cyclic GMP/antagonists & inhibitors , Deuteroporphyrins/metabolism , Heme Oxygenase (Decyclizing)/antagonists & inhibitors , Heme/analogs & derivatives , Heme/metabolism , Lysine/analogs & derivatives , Lysine/metabolism , Nociceptive Pain/metabolism , Oxadiazoles/pharmacology , Pain Measurement/methods , Rats, Wistar , Signal Transduction/physiology
8.
Braz. j. med. biol. res ; 47(1): 42-49, 01/2014. tab, graf
Article in English | LILACS | ID: lil-697672

ABSTRACT

Membranous nephropathy (MN), characterized by the presence of diffuse thickening of the glomerular basement membrane and subepithelial in situ immune complex disposition, is the most common cause of idiopathic nephrotic syndrome in adults, with an incidence of 5-10 per million per year. A number of studies have confirmed the relevance of several experimental insights to the pathogenesis of human MN, but the specific biomarkers of MN have not been fully elucidated. As a result, our knowledge of the alterations in histone methylation in MN is unclear. We used chromatin immunoprecipitation followed by high-throughput sequencing (ChIP-seq) to analyze the variations in a methylated histone (H3K9me3) in peripheral blood mononuclear cells from 10 MN patients and 10 healthy subjects. There were 108 genes with significantly different expression in the MN patients compared with the normal controls. In MN patients, significantly increased activity was seen in 75 H3K9me3 genes, and decreased activity was seen in 33, compared with healthy subjects. Five positive genes, DiGeorge syndrome critical region gene 6 (DGCR6), sorting nexin 16 (SNX16), contactin 4 (CNTN4), baculoviral IAP repeat containing 3 (BIRC3), and baculoviral IAP repeat containing 2 (BIRC2), were selected and quantified. There were alterations of H3K9me3 in MN patients. These may be candidates to help explain pathogenesis in MN patients. Such novel findings show that H3K9me3 may be a potential biomarker or promising target for epigenetic-based MN therapies.


Subject(s)
Adult , Female , Humans , Male , Glomerulonephritis, Membranous/genetics , Histones/genetics , Leukocytes, Mononuclear/metabolism , Lysine/genetics , Case-Control Studies , Chromatin Immunoprecipitation , Glomerulonephritis, Membranous/metabolism , Histones/metabolism , Lysine/metabolism , Methylation
9.
Yonsei Medical Journal ; : 377-385, 2012.
Article in English | WPRIM | ID: wpr-154804

ABSTRACT

PURPOSE: The purpose of the present study was to investigate the aberrance of histone H3 lysine 4 trimethylation (H3K4me3) in patients with IgA Nephropathy (IgAN). MATERIALS AND METHODS: In this study, H3K4me3 variations in peripheral blood mononuclear cells (PBMCs) from 15 IgAN patients and 15 healthy subjects were analyzed using chromatin immunoprecipitation linked to microarrays analysis (ChIP-chip). ChIP real-time PCR was used to validate the microarray results. Expression analysis by quantitative real-time PCR (qRT-PCR) revealed correlations between mRNA and H3K4me3 levels. DNA methylation status was analyzed by quantitative methylation-specific PCR. RESULTS: We found that 321 probes displayed significant H3K4me3 differences in IgAN patients compared with healthy controls. Among these probes, 154 probes displayed increased H3K4me3 and 167 probes demonstrated decreased H3K4me3. For further validation, we selected 4 key relevant genes (FCRL4, GALK2, PTPRN2 and IL1RAPL1) to study. The results of ChIP real-time PCR coincided well with the microarray data. Quantitative RT-PCR revealed the correlations between the mRNA expression and the methylation levels of H3K4me3. Different degrees of DNA methylation alterations appeared on the selected positive genes. CONCLUSION: Our studies indicated that there were significant alterations in H3K4me3 in IgAN patients. These findings may help to explain the disturbed immunity and abnormal glycosylation involved in IgAN patients.


Subject(s)
Adult , Case-Control Studies , Chromatin Immunoprecipitation , Female , Glomerulonephritis, IGA/genetics , Histones/metabolism , Humans , Leukocytes, Mononuclear/metabolism , Lysine/metabolism , Male , Methylation , Oligonucleotide Array Sequence Analysis/methods , Real-Time Polymerase Chain Reaction , Young Adult
10.
Article in English | WPRIM | ID: wpr-214630

ABSTRACT

Epigenomic and genomic changes affect gene expression and contribute to tumor development. The histone modifications trimethylated histone H3 lysine 4 (H3K4me3) and lysine 27 (H3K27me3) are epigenetic regulators associated to active and silenced genes, respectively and alterations of these modifications have been observed in cancer. Furthermore, genomic aberrations such as DNA copy number changes are common events in tumors. Pheochromocytoma is a rare endocrine tumor of the adrenal gland that mostly occurs sporadic with unknown epigenetic/genetic cause. The majority of cases are benign. Here we aimed to combine the genome-wide profiling of H3K4me3 and H3K27me3, obtained by the ChIP-chip methodology, and DNA copy number data with global gene expression examination in a malignant pheochromocytoma sample. The integrated analysis of the tumor expression levels, in relation to normal adrenal medulla, indicated that either histone modifications or chromosomal alterations, or both, have great impact on the expression of a substantial fraction of the genes in the investigated sample. Candidate tumor suppressor genes identified with decreased expression, a H3K27me3 mark and/or in regions of deletion were for instance TGIF1, DSC3, TNFRSF10B, RASSF2, HOXA9, PTPRE and CDH11. More genes were found with increased expression, a H3K4me3 mark, and/or in regions of gain. Potential oncogenes detected among those were GNAS, INSM1, DOK5, ETV1, RET, NTRK1, IGF2, and the H3K27 trimethylase gene EZH2. Our approach to associate histone methylations and DNA copy number changes to gene expression revealed apparent impact on global gene transcription, and enabled the identification of candidate tumor genes for further exploration.


Subject(s)
Adrenal Gland Neoplasms/genetics , Epigenesis, Genetic , Female , Gene Dosage/genetics , Gene Expression Regulation, Neoplastic , Gene Regulatory Networks/genetics , Genome, Human/genetics , Genomics , Histones/metabolism , Humans , Lysine/metabolism , Methylation , Pheochromocytoma/genetics , Protein Processing, Post-Translational , Tumor Suppressor Proteins/genetics
11.
J Genet ; 2008 Dec; 87(3): 235-40
Article in English | IMSEAR | ID: sea-114483

ABSTRACT

In Drosophila melanogaster, dosage compensation occurs through hypertranscription of sex-linked genes in males. The hypertranscription involves acetylation of histone 4 at lysine 16 (H4K16) on amale X-chromosome, brought about by a histone acetyltransferase encoded by the dosage compensation gene, males absent on the first (mof). We report a phenomenon in the strain In(1)B(M2)(reinverted) of D. melanogaster where the global structure of the male X-chromosome can be altered at the third instar larval stage through a 4-h cold shock at 12+/-1 degrees C. We show that the cold shock results in a transient hyperacetylation of H4K16 and an increased expression of MOF. Control proteins H4 acetylated at lysine 5, and the dosage compensation gene msl-2, do not show any change in expression after cold shock. Cytology of the male X-chromosome at different time points during cold shock and recovery, suggests that the hyperacetylation of H4 at lysine 16 causes the X-chromosome to corkscrew into itself, thereby achieving the cold-induced change in the higher order structure of the male polytene X-chromosome. Our studies suggest a role for H4K16 in maintaining the structure of the male X-chromosome in Drosophila.


Subject(s)
Acetylation , Animals , Cell Nucleus/metabolism , Cold Temperature , Drosophila melanogaster/metabolism , Female , Histones/metabolism , Immunoblotting , Larva/metabolism , Lysine/metabolism , Male , X Chromosome/metabolism
12.
Medical Principles and Practice. 2008; 17 (3): 258-261
in English | IMEMR | ID: emr-104587

ABSTRACT

To report the importance of a rare organic acid metabolic disorder, L-2-hydroxyglutaric aciduria, and its characteristic neuroimaging cerebral white matter abnormalities in a case of epilepsy. A 19-year-old male presented with an 11-year history consisting of school failures, intellectual deterioration and generalized tonic-clonic convulsions. Neurological examination showed mental subnormality, mild dysarthria and bilateral pyramidal signs. Computed tomography and magnetic resonance imaging [MRI] of the brain showed characteristic white matter lesions, suggestive of L-2-hydroxyglutaric aciduria. The diagnosis of this disease was confirmed by elevated urinary concentrations of L-2-hydroxyglutaric acid. The epilepsy was partially controlled with antiepileptic drugs. This report indicates the importance of routine examination of urinary organic acids in children and young adults presenting with chronic encephalopathy and epilepsy with characteristic MRI white matter lesions. L-2-hydroxyglutaric aciduria should be considered as one of the differential diagnoses of epilepsy


Subject(s)
Humans , Male , Epilepsy, Tonic-Clonic/urine , Epilepsy, Tonic-Clonic/physiopathology , Alcohol Oxidoreductases/urine , Biomarkers/urine , Risk Factors , Lysine/metabolism , Brain Diseases, Metabolic/diagnosis , Magnetic Resonance Imaging , Electroencephalography , Cerebrospinal Fluid
13.
Article in English | WPRIM | ID: wpr-37552

ABSTRACT

Various cell types in higher multicellular organisms are genetically homogenous, but are functionally and morphologically heterogeneous due to the differential expression of genes during development, which appears to be controlled by epigenetic mechanisms. However, the exact molecular mechanisms that govern the tissue-specific gene expression are poorly understood. Here, we show that dynamic changes in histone modifications and DNA methylation in the upstream coding region of a gene containing the transcription initiation site determine the tissue-specific gene expression pattern. The tissue-specific expression of the transgene correlated with DNA demethylation at specific CpG sites as well as significant changes in histone modifications from a low ratio of methylated H3- lysine 4 or acetylated H3-lysine 9, 14 to acetylated H4 to higher ratios. Based on the programmed status of transgene silenced in cloned mammalian ear-derived fibroblasts, the transgene could be reprogrammed by change of histone modification and DNA methylation by inhibiting both histone deacetylase and DNA methylation, resulting in high expression of the transgene. These findings indicate that dynamic change of histone modification and DNA methylation is potentially important in the establishment and maintenance of tissue-specific gene expression.


Subject(s)
Transgenes/genetics , Swine , Organ Specificity/genetics , Methylation , Lysine/metabolism , Histones/metabolism , Histone Deacetylases/metabolism , Gene Silencing , Gene Expression , Fibroblasts , Ear , DNA Methylation , Cells, Cultured , Animals, Genetically Modified , Animals , Acetylation
14.
Article in English | WPRIM | ID: wpr-201948

ABSTRACT

The methylation of a 23-kDa nuclear protein increased after partial hepatectomy and methylation returned to basal levels after the initial stage of regeneration. The methylating enzyme was partially purified from rat liver by ammonium sulfate precipitation, DEAE-anion exchange chromatography and Butyl-Sepharose chromatography. The 23-kDa protein was purified from a nuclear fraction of liver tissue with SP-Sepharose. When the 23-kDa protein was methylated with the partially purified methyltransferase and analyzed on C18 high performance liquid chromatography (HPLC), the methylated acceptor amino acid was monomethyl lysine (MML). Previously, only arginine N-methylation of specific substrate proteins has been reported during liver regeneration. However, in this report, we found that lysine N-methylation increased during early hepatic regeneration, suggesting that lysine N-methylation of the 23-kDa nuclear protein may play a functional role in hepatic regeneration. The methyltransferase did not methylate other proteins such as histones, hnRNPA1, or cytochrome C, suggesting the enzyme is a 23-kDa nuclear protein- specific lysine N-methyltransferase.


Subject(s)
Animals , Cytochromes c/metabolism , DNA Helicases/metabolism , Hepatectomy , Histone-Lysine N-Methyltransferase/metabolism , Histones/metabolism , Liver , Liver Regeneration/physiology , Lysine/metabolism , Methylation , Proteins/metabolism , Rats , Rats, Sprague-Dawley
15.
J Biosci ; 2003 Sep; 28(5): 597-604
Article in English | IMSEAR | ID: sea-111002

ABSTRACT

Twenty soil communities from the northeastern forests (Assam) and the Western Ghats (Maharashtra) were screened for the presence of the lysine aminotransferase (lat) gene from Nocardia. Hybridization probes and primers were synthesized in accordance with the reported sequence of the Nocardia lat gene from GenBank (number: G1 49355). Seven positives were obtained from the 20 soils. Six of the seven positive were from the Western Ghats and one from the northeast Assam forests. Eighteen actinomycete isolates from the 7 positive soils showed the presence of the lat gene. Only 9 isolates actually produced an antibiotic. These results are discussed.


Subject(s)
Bacterial Proteins/genetics , India , Lysine/metabolism , Molecular Sequence Data , Nocardia/enzymology , Soil Microbiology , Transaminases/genetics
16.
Article in English | WPRIM | ID: wpr-13862

ABSTRACT

Transglutaminase enzymes (TGases) catalyze the calcium dependent formation of an isopeptide bond between protein-bound glutamine and lysine substrates. Previously we have shown that activated TGase 3 acquires two additional calcium ions at site two and three. The calcium ion at site three results in the opening of a channel. At this site, the channel opening and closing could modulate, depending on which metal is bound. Here we propose that the front of the channel could be used by the two substrates for enzyme reaction. We propose that the glutamine substrate is directed from Trp236 into the enzyme, shown by molecular docking. Then a lysine substrate approaches the opened active site to engage Trp327, leading to formation of the isopeptide bond. Further, direct comparisons of the structures of TGase 3 with other TGases have allowed us to identify several residues that might potentially be involved in generic and specific recognition of the glutamine and lysine substrates.


Subject(s)
Animals , Binding Sites , Calcium/metabolism , Calcium-Binding Proteins/metabolism , Glutamine/metabolism , Humans , Lysine/metabolism , Models, Chemical , Models, Molecular , Protein Binding , Protein Structure, Tertiary , Transglutaminases/metabolism
17.
Article in English | IMSEAR | ID: sea-21984

ABSTRACT

BACKGROUND & OBJECTIVES: Studies have been carried out to synthesize and characterize the photoconjugate between positively charged amino acid, arginine and DNA fragments and their role in the induction of anti-DNA antibodies. METHODS: Calf thymus DNA fragments of about 200 base pairs (bp) were covalently crosslinked with arginine under UV light. The amino acid was found to be covalently photoconjugated to DNA and resulted in the formation of a crosslink. The photoadduct was characterized by various physicochemical methods. RESULTS: Photoaddition of arginine to 200 bp DNA rendered the nucleic acid conformer thermodynamically more stable than the native form. After systematic characterization of the photoadduct, it was used as an antigen for the generation of antibodies in experimental animals. The photoadducts were found to be immunogenic in rabbits, inducing high titre antibodies. The DNA-arginine photoadduct showed higher binding with SLE sera known to have high level of anti-DNA antibodies. INTERPRETATION & CONCLUSION: Naturally occurring anti-DNA autoantibodies were found to recognize DNA-arginine photoadduct. The recognition of DNA-arginine photoadduct by anti-DNA autoantibodies points to the role of modified DNA in the induction of anti-DNA antibodies in autoimmune disorders.


Subject(s)
Animals , Antibodies, Antinuclear/immunology , Arginine/metabolism , Autoantibodies/immunology , DNA Adducts/immunology , Humans , Light , Lupus Erythematosus, Systemic/immunology , Lysine/metabolism , Nucleic Acid Conformation , Rabbits
18.
Medicina (B.Aires) ; 60(5/1): 580-6, 2000. graf
Article in Spanish | LILACS | ID: lil-275468

ABSTRACT

El clonixinato de lisina (CL) es un antiinflamatorio no esteroide (AINE) con pocos efectos adversos, por lo que se ha postulado que a concentraciones equivalentes a las encontradas en plasma humano después de dosis terapéuticas inhibiría en escaso grado la cicloxigenasa I (COX-I). Se efectuaron 3 experimentos. Experimento 1: se estudió el efecto in vitro de CL en concentraciones de 4 y 6 ug/ml, las que se corresponden con las alcanzadas en plasma con una dosis oral de 125 mg. Los segmentos de vesícula biliar (n = 6) se incubaron con 0.25 uCi de ácido araquidónico 14C y se midió la producción de prostaglandina E 2, prostaglandina F 2a y prostaglandina 6 ceto F 1a. El CL no modificó la producción basal de ninguna de las tres prostaglandinas, pero con 6 ug/ml disminuyó significativamente la producción de ácido 5- hidroxieicosatetraenico (5-HETE). Experimento 2: se administró una infusión continua de CL a 6 pacientes en el pre operatorio inmediato para lograr una concentración en estado estacioanrio entre 4 y 6 ug/ml. Se incubaron segmentos de vesícula biliar de estos 6 pacientes y de 6 pacientes control no tratados con ácido araquidónico 14 C. Se observó que los segmentos de vesícula biliar tratados con CL no mostraron inhibición de la producción de ninguna de las tres PGs, mientras que el 5-HETE liberado al medio fue significativamente menor. Experimento 3: 18 pacientes recibieron bolos EV de: CL 100mg (n1, = 6); CL 200 mg (n2, = 6) o indometacina (INDO) 50 mg (n3 = 6). Con ninguna de las dos dosis de CL se obtuvo inhibición de la síntesis de PGs, por el contrario de INDO inhibió su síntesis. Cuando se valoró la producción de 5-HETE, los dos AINES estudiados de se comportaron en forma distinta. El tratamiento con INDO no modificó el 5-F-HETE producido, mientras que el tratamiento con CL lo inhibió significativamente. En los tres tipos de estudios realizados in vitro e in vivo: infusión continua y bolo IV, el CL no inhibió la síntesis de PGs y disminuyó significativamente el 5-HETE.


Subject(s)
Humans , Anti-Inflammatory Agents, Non-Steroidal/pharmacology , Gallbladder/drug effects , Hydroxyeicosatetraenoic Acids/biosynthesis , Lysine/analogs & derivatives , Lysine/pharmacology , Prostaglandins/biosynthesis , Anti-Inflammatory Agents, Non-Steroidal/metabolism , Blotting, Western , Gallbladder/enzymology , Indomethacin/pharmacology , Lysine/metabolism
19.
Indian J Biochem Biophys ; 1996 Dec; 33(6): 519-22
Article in English | IMSEAR | ID: sea-28963

ABSTRACT

Isatin (2,3-dioxoindole) competitively inhibited (27-40%) Na(+)-dependent L-lysine uptake in rat intestine. The value of Kt was increased from 3.04 mM in control to 5.88 mM in presence of 10 mM isatin. Effect of isatin on the Na(+)-independent amino acid uptake was insignificant (12-18%). The inhibitory constant (Ki) was 2.8 mM under these conditions. The observed inhibition was unaffected by -SH group reacting agents. Isatin (1-10 mM) inhibited Na+, K(+)-ATPase activity in intestine in vitro, the maximum inhibition (66%) being at 10 mM isatin concentration. But the drug had no effect on enzyme activity under in vivo conditions.


Subject(s)
Animals , Biological Transport/drug effects , Intestinal Mucosa/metabolism , Isatin/pharmacology , Kinetics , Lysine/metabolism , Rats , Sodium/pharmacology , Sodium-Potassium-Exchanging ATPase/antagonists & inhibitors , Sulfhydryl Reagents/pharmacology
20.
Vet. Méx ; 24(4): 311-6, oct.-dic. 1993. ilus, tab
Article in Spanish | LILACS | ID: lil-139053

ABSTRACT

Se efectuaron dos experimentos para observar la respuesta de pollos de engorda en la etapa de iniciación, alimentados con dietas con diferentes niveles de energía, proteína y lisina. El primero se realizó bajo un sideño "San Cristóbal" de superficie de respuesta. Se utilizaron 480 pollos de siete días de edad, distribuidos al azar en 12 tratamientos. Los resultados, después de 21 días de experimentación, indicaron que los aumentos de peso de los pollos están influenciados lineal y significativamente (P < 0.01) por el nivel de energía de la dieta. El consumo de alimento estuvo influenciado (P < 0.05) por el nivel de lisina y proteína, pero no por el nivel de energía. Por lo que respecta a conversión alimenticia, se encontraron diferencias (P < 0.01) entre tratamientos debido a proteína, lisina y energía. En el segundo experimento, se evaluó la respuesta de los pollos (0-28 días) alimentados con dietas con diferntes niveles de proteína y lisina con 3000 Kcal de EM/kg. Los resultados indicaron que los niveles de proteína no tuvieron ningún efecto en ganancia de peso (P > 0.05), pero que al aumentar la lisina mejoró la ganancia de peso (P < 0.01). En consumo de alimento, se encontraron diferencias significativas entre tratamientos. En conversión alimenticia, no existieron diferencias (P > 0.05) entre los diversos niveles de proteína, aunque sí para lisina


Subject(s)
Animals , Animal Feed/analysis , Animal Feed/supply & distribution , Chickens/growth & development , Chickens/metabolism , Diet Therapy , Diet Therapy/veterinary , Lysine/metabolism , Weight Gain
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