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1.
BMC Biotechnol ; 22(1): 30, 2022 10 27.
Artigo em Inglês | MEDLINE | ID: mdl-36303174

RESUMO

BACKGROUND: An important conceptual advance in health and the environment has been recognized that enzymes play a key role in the green processing industries. Of particular interest, chitosanase is beneficial for recycling the chitosan resource and producing chitosan oligosaccharides. Also, chitosan gene expression and molecular characterization will promote understanding of the biological function of bacterial chitosanase as well as explore chitosanase for utilizing chitosan resources. RESULTS: A chitosanase-producing bacterium TY24 was isolated and identified as Bacillus cereus. Moreover, the chitosanase gene was cloned and expressed in Escherichia coli. Sequence analysis reveals that the recombinant chitosanase (CHOE) belongs to the glycoside hydrolases 8 family. The purified CHOE has a molecular weight of about 48 kDa and the specific activity of 1150 U/mg. The optimal pH and temperature of CHOE were 5.5 and 65 °C, respectively. The enzyme was observed stable at the pH range of 4.5-7.5 and the temperature range of 30-65 °C. Especially, the half-life of CHOE at 65 °C was 161 min. Additionally, the activity of CHOE was remarkably enhanced in the presence of Mn2+, Cu2+, Mg2+ and K+, beside Ca2+ at 5 mM. Especially, the activity of CHOE was enhanced to more than 120% in the presence of 1% of various surfactants. CHOE exhibited the highest substrate specificity toward colloid chitosan. CONCLUSION: A bacterial chitosanase was cloned from B. cereus and successfully expressed in E. coli (BL21) DE3. The recombinant enzyme displayed good stability under acid pH and high-temperature conditions.


Assuntos
Bacillus cereus , Quitosana , Bacillus cereus/genética , Bacillus cereus/metabolismo , Quitosana/metabolismo , Escherichia coli/genética , Escherichia coli/metabolismo , Glicosídeo Hidrolases/química , Clonagem Molecular , Concentração de Íons de Hidrogênio
2.
Neoplasma ; 68(6): 1301-1309, 2021 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-34648299

RESUMO

This study aimed to measure the expression of SAA2 in plasma and to assess its diagnostic efficacy as a biomarker for non-small cell lung cancer (NSCLC). The gene expression of SAA2 in NSCLC was analyzed based on a database. Then, SAA2 expression was detected by immunohistochemistry in lung tissue and by enzyme-linked immunosorbent assay in 90 patients with NSCLC and 61 normal controls. Finally, the diagnostic performance was assessed in terms of accuracy, sensitivity, and specificity. At the gene and protein levels, the SAA2 expression was significantly higher in the NSCLC group than in the control group (p<0.01). It was higher in lung squamous carcinoma than in lung adenocarcinoma and in males than in females, and this trend was also observed in the lung squamous carcinoma group. Of note, the expression of SAA2 increased with increasing disease stage. Receiver operating characteristic (ROC) curve analysis revealed that the sensitivity of SAA2 was 83.61%, the specificity was 91.11%, and the area under the curve (AUC) was 0.9252. Its accuracy was 68.89%, which was higher than that of other conventional diagnostic biomarkers, and the combined application can effectively improve the diagnostic efficiency. Based on the results, SAA2 expression was positively correlated with the disease stage of NSCLC. Notably, SAA2 is more concerning in male patients with lung squamous carcinoma, and it can help in the screening and diagnosis of NSCLC. SAA2 may represent a novel diagnostic biomarker in NSCLC.


Assuntos
Carcinoma Pulmonar de Células não Pequenas , Neoplasias Pulmonares , Área Sob a Curva , Biomarcadores Tumorais/genética , Carcinoma Pulmonar de Células não Pequenas/diagnóstico , Carcinoma Pulmonar de Células não Pequenas/genética , Feminino , Humanos , Neoplasias Pulmonares/diagnóstico , Neoplasias Pulmonares/genética , Masculino , Curva ROC , Proteína Amiloide A Sérica/genética
3.
Huan Jing Ke Xue ; 40(5): 2094-2100, 2019 May 08.
Artigo em Chinês | MEDLINE | ID: mdl-31087845

RESUMO

To learn about the status of antibiotic contamination and their ecological risks in Chinese surface-water environments, the risk quotient (RQ) and joint risk quotient (RQSUM) methods were applied to assess the ecological risks of five typical surface-water environments in China during the flood season. The results showed that the main types of antibiotic contamination in the five regions were sulfamethoxazole (SMX), sulfamethazine (SM), erythromycin (ETM), roxithromycin (RTM), tetracycline (TC), oxytetracycline (OTC), norfloxacin (NOR), and ofloxacin (OFL). Among eight types of antibiotic contamination, sulfamethoxazole (SMX) and erythromycin (ETM) occupied a dominant position. The contribution rate of SMX in the Yangtze River Delta and Chaohu Basin was 91.1% and 98.5%, respectively. Meanwhile, the contribution rates of ETM in Jianghan Plain, Pearl River Delta, and Yellow River Delta were 94.4%, 81.8%, and 60%, respectively. Based on the joint risk quotients (RQSUM), the order of ecological risks in the research areas was:Jianghan Plain (20.204) > Yangtze River Delta (8.769) > Chaohu Basin (2.692) > Yellow River Delta (1.943) > Pearl River Delta (1.222).


Assuntos
Antibacterianos/análise , Monitoramento Ambiental , Poluentes Químicos da Água/análise , China , Medição de Risco , Rios
4.
BMC Dev Biol ; 18(1): 20, 2018 11 20.
Artigo em Inglês | MEDLINE | ID: mdl-30458702

RESUMO

BACKGROUND: Histone modifications are critical in regulating neuronal processes. However, the impacts of individual histone modifications on learning and memory are elusive. Here, we investigated the contributions of histone H3 lysine modifications to learning and memory in Drosophila by using histone lysine-to-alanine mutants. RESULTS: Behavioural analysis indicated that compared to the H3WT group, mutants overexpressing H3K23A displayed impaired courtship learning. Chromatin immunoprecipitation analysis of H3K23A mutants showed that H3K23 acetylation (H3K23ac) levels were decreased on learning-related genes. Knockdown of CREB-binding protein (CBP) decreased H3K23ac levels, attenuated the expression of learning-related genes, led to a courtship learning defect and altered development of the mushroom bodies. A decline in courtship learning ability was observed in both larvae and adult treatments with ICG-001. Furthermore, treatment of Drosophila overexpressing mutated H3K23A with a CBP inhibitor did not aggravate the learning defect. CONCLUSIONS: H3K23ac, catalysed by the acetyltransferases dCBP, contributes to Drosophila learning, likely by controlling the expression of specific genes. This is a novel epigenetic regulatory mechanism underlying neuronal behaviours.


Assuntos
Proteínas de Drosophila/metabolismo , Drosophila melanogaster/metabolismo , Histonas/metabolismo , Lisina/metabolismo , Fatores de Transcrição de p300-CBP/metabolismo , Acetilação , Animais , Corte , Feminino , Regulação da Expressão Gênica , Técnicas de Silenciamento de Genes , Histonas/genética , Aprendizagem , Masculino , Mutação , Neurônios/metabolismo
5.
J Cell Sci ; 131(12)2018 06 21.
Artigo em Inglês | MEDLINE | ID: mdl-29760279

RESUMO

Dysregulation of the homeostatic balance of histone H3 di- and tri-methyl lysine 27 (H3K27me2/3) levels caused by the mis-sense mutation of histone H3 (H3K27M) is reported to be associated with various types of cancers. In this study, we found that reduction in H3K27me2/3 caused by H3.1K27M, a mutation of H3 variants found in patients with diffuse intrinsic pontine glioma (DIPG), dramatically attenuated the presence of 53BP1 (also known as TP53BP1) foci and the capability of non-homologous end joining (NHEJ) in human dermal fibroblasts. H3.1K27M mutant cells showed increased rates of genomic insertions/deletions and copy number variations, as well as an increase in p53-dependent apoptosis. We further showed that both hypo-H3K27me2/3 and H3.1K27M interacted with FANCD2, a central player in the choice of DNA repair pathway. H3.1K27M triggered the accumulation of FANCD2 on chromatin, suggesting an interaction between H3.1K27M and FANCD2. Interestingly, knockdown of FANCD2 in H3.1K27M cells recovered the number of 53BP1-positive foci, NHEJ efficiency and apoptosis rate. Although these findings in HDF cells may differ from the endogenous regulation of the H3.1K27M mutant in the specific tumor context of DIPG, our results suggest a new model by which H3K27me2/3 facilitates NHEJ and the maintenance of genome stability.This article has an associated First Person interview with the first author of the paper.


Assuntos
Cromatina/metabolismo , Reparo do DNA por Junção de Extremidades , Enzimas Reparadoras do DNA/metabolismo , Proteínas de Ligação a DNA/metabolismo , Proteína do Grupo de Complementação D2 da Anemia de Fanconi/metabolismo , Histonas/metabolismo , Neoplasias do Tronco Encefálico/genética , Neoplasias do Tronco Encefálico/metabolismo , Linhagem Celular , Cromatina/genética , Reparo do DNA , Enzimas Reparadoras do DNA/genética , Proteínas de Ligação a DNA/genética , Proteína do Grupo de Complementação D2 da Anemia de Fanconi/genética , Fibroblastos , Instabilidade Genômica , Glioma/genética , Glioma/metabolismo , Células HEK293 , Histonas/genética , Humanos , Metilação , Proteína 1 de Ligação à Proteína Supressora de Tumor p53/genética , Proteína 1 de Ligação à Proteína Supressora de Tumor p53/metabolismo
6.
Nat Commun ; 6: 8856, 2015 Nov 19.
Artigo em Inglês | MEDLINE | ID: mdl-26581759

RESUMO

Epigenetics plays critical roles in controlling stem cell self-renewal and differentiation. Histone H1 is one of the most critical chromatin regulators, but its role in adult stem cell regulation remains unclear. Here we report that H1 is intrinsically required in the regulation of germline stem cells (GSCs) in the Drosophila ovary. The loss of H1 from GSCs causes their premature differentiation through activation of the key GSC differentiation factor bam. Interestingly, the acetylated H4 lysine 16 (H4K16ac) is selectively augmented in the H1-depleted GSCs. Furthermore, overexpression of mof reduces H1 association on chromatin. In contrast, the knocking down of mof significantly rescues the GSC loss phenotype. Taken together, these results suggest that H1 functions intrinsically to promote GSC self-renewal by antagonizing MOF function. Since H1 and H4K16 acetylation are highly conserved from fly to human, the findings from this study might be applicable to stem cells in other systems.


Assuntos
Autorrenovação Celular , Proteínas de Drosophila/metabolismo , Drosophila melanogaster/metabolismo , Células Germinativas/metabolismo , Histonas/metabolismo , Motivos de Aminoácidos , Animais , Proteínas de Drosophila/química , Proteínas de Drosophila/genética , Drosophila melanogaster/química , Drosophila melanogaster/genética , Epigênese Genética , Feminino , Células Germinativas/citologia , Histonas/química , Histonas/genética , Masculino , Ovário/química , Ovário/metabolismo
7.
Proc Natl Acad Sci U S A ; 112(45): 13988-93, 2015 Nov 10.
Artigo em Inglês | MEDLINE | ID: mdl-26508632

RESUMO

Dynamic regulation of chromatin structure is required to modulate the transcription of genes in eukaryotes. However, the factors that contribute to the plasticity of heterochromatin structure are elusive. Here, we report that cyclin-dependent kinase 12 (CDK12), a transcription elongation-associated RNA polymerase II (RNAPII) kinase, antagonizes heterochromatin enrichment in Drosophila chromosomes. Notably, loss of CDK12 induces the ectopic accumulation of heterochromatin protein 1 (HP1) on euchromatic arms, with a prominent enrichment on the X chromosome. Furthermore, ChIP and sequencing analysis reveals that the heterochromatin enrichment on the X chromosome mainly occurs within long genes involved in neuronal functions. Consequently, heterochromatin enrichment reduces the transcription of neuronal genes in the adult brain and results in a defect in Drosophila courtship learning. Taken together, these results define a previously unidentified role of CDK12 in controlling the epigenetic transition between euchromatin and heterochromatin and suggest a chromatin regulatory mechanism in neuronal behaviors.


Assuntos
Montagem e Desmontagem da Cromatina/fisiologia , Quinases Ciclina-Dependentes/metabolismo , Drosophila/genética , Epigênese Genética/fisiologia , Heterocromatina/fisiologia , Aprendizagem/fisiologia , Animais , Sequência de Bases , Western Blotting , Montagem e Desmontagem da Cromatina/genética , Imunoprecipitação da Cromatina , Drosophila/fisiologia , Heterocromatina/genética , Imunoprecipitação , Dados de Sequência Molecular , Octoxinol , Reação em Cadeia da Polimerase em Tempo Real , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Glândulas Salivares/anatomia & histologia , Glândulas Salivares/metabolismo , Alinhamento de Sequência , Análise de Sequência de DNA
8.
J Cell Sci ; 125(Pt 22): 5369-78, 2012 Nov 15.
Artigo em Inglês | MEDLINE | ID: mdl-22956542

RESUMO

Core histone modifications play an important role in chromatin remodeling and transcriptional regulation. Histone acetylation is one of the best-studied gene modifications and has been shown to be involved in numerous important biological processes. Herein, we demonstrated that the depletion of histone deacetylase 3 (Hdac3) in Drosophila melanogaster resulted in a reduction in body size. Further genetic studies showed that Hdac3 counteracted the organ overgrowth induced by overexpression of insulin receptor (InR), phosphoinositide 3-kinase (PI3K) or S6 kinase (S6K), and the growth regulation by Hdac3 was mediated through the deacetylation of histone H4 at lysine 16 (H4K16). Consistently, the alterations of H4K16 acetylation (H4K16ac) induced by the overexpression or depletion of males-absent-on-the-first (MOF), a histone acetyltransferase that specifically targets H4K16, resulted in changes in body size. Furthermore, we found that H4K16ac was modulated by PI3K signaling cascades. The activation of the PI3K pathway caused a reduction in H4K16ac, whereas the inactivation of the PI3K pathway resulted in an increase in H4K16ac. The increase in H4K16ac by the depletion of Hdac3 counteracted the PI3K-induced tissue overgrowth and PI3K-mediated alterations in the transcription profile. Overall, our studies indicated that Hdac3 served as an important regulator of the PI3K pathway and revealed a novel link between histone acetylation and growth control.


Assuntos
Proteínas de Drosophila/metabolismo , Drosophila melanogaster/enzimologia , Drosophila melanogaster/crescimento & desenvolvimento , Histona Desacetilases/metabolismo , Histonas/metabolismo , Lisina/metabolismo , Especificidade de Órgãos , Fosfatidilinositol 3-Quinases/metabolismo , Acetilação , Animais , Tamanho Corporal , Tamanho Celular , Proteínas de Drosophila/deficiência , Drosophila melanogaster/citologia , Drosophila melanogaster/ultraestrutura , Feminino , Histona Acetiltransferases/metabolismo , Histona Desacetilases/deficiência , Insulina/metabolismo , Masculino , Proteínas Nucleares/metabolismo , Receptor de Insulina/metabolismo , Proteínas Quinases S6 Ribossômicas/metabolismo , Transdução de Sinais , Transcrição Gênica
9.
J Biol Chem ; 286(11): 9020-30, 2011 Mar 18.
Artigo em Inglês | MEDLINE | ID: mdl-21205821

RESUMO

The turnover of tumor suppressor p53 is critical for its role in various cellular events. However, the pathway that regulates the turnover of the Drosophila melanogaster DMP53 is largely unknown. Here, we provide evidence for the first time that the E2 ligase, Drosophila homolog of Rad6 (dRad6/Dhr6), plays an important role in the regulation of DMP53 turnover. Depletion of dRad6 results in DMP53 accumulation, whereas overexpression of dRad6 causes enhanced DMP53 degradation. We show that dRad6 specifically interacts with DMP53 at the transcriptional activation domain and regulates DMP53 ubiquitination. Loss of dRad6 function in transgenic flies leads to lethalities and altered morphogenesis. The dRad6-induced defects in cell proliferation and apoptosis are found to be DMP53-dependent. The loss of dRad6 induces an accumulation of DMP53 that enhances the activation of apoptotic genes and leads to apoptosis in the presence of stress stimuli. In contrast to that, the E3 ligase is the primary factor that regulates p53 turnover in mammals, and this work demonstrates that the E2 ligase dRad6 is critical for the control of DMP53 degradation in Drosophila.


Assuntos
Proteínas Reguladoras de Apoptose/metabolismo , Apoptose/fisiologia , Proteínas de Drosophila/metabolismo , Proteína Supressora de Tumor p53/metabolismo , Enzimas de Conjugação de Ubiquitina/metabolismo , Animais , Proteínas Reguladoras de Apoptose/genética , Linhagem Celular , Proteínas de Drosophila/genética , Drosophila melanogaster , Morfogênese/fisiologia , Estrutura Terciária de Proteína , Proteína Supressora de Tumor p53/genética , Enzimas de Conjugação de Ubiquitina/genética
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