AIM To evaluate the relationship between serum adenosine deaminase(ADA) levels and histological features in patients with autoimmune hepatitis(AIH).METHODS A total of 80 subjects(52 AIH cases and 28 healthy controls) ...AIM To evaluate the relationship between serum adenosine deaminase(ADA) levels and histological features in patients with autoimmune hepatitis(AIH).METHODS A total of 80 subjects(52 AIH cases and 28 healthy controls) were included in the study. Patients were diagnosed according to the simplified criteria suggested by the International Autoimmune Hepatitis Group. All of the cases had been diagnosed with AIH between 2010-2015 at Hacettepe University, Department of Gastroenterology. Serum blood samples were collected and stored at-80 ℃ until the biochemical estimation of ADA activity. The diagnosis of patients was confirmed by liver biopsy. Serum ADA > 20 U/L was considered to be high level.RESULTS Mean serum ADA levels were significantly higher in AIH patients than those in healthy controls(25.4 ± 9.6 U/L vs 12.8 ± 2.2 U/L, P < 0.001). Serum ADA levels > 20 U/L were found in 63.5% AIH patients and in 0% healthy controls(P < 0.001). Mean serum ADA levels were significantly increased in each stage of histological activity: 15.2 ± 3.5 U/L for patients with mild interface hepatitis, 23.1 ± 10.0 U/L for patients with moderate interface hepatitis and 30.9 ± 7.0 U/L for patients with severe interface hepatitis(P < 0.001). Correlation analysis showed that there was a positive association between serum ADA levels and histological activity(r = 0.71, P < 0.001). Receiver operating characteristic analysis suggested that 24.5 U/L was the optimum cut-off point of ADA level for severe interface hepatitis(sensitivity 88%, specificity 85.2%, area under thecurve: 0.88).CONCLUSION Because of the positive correlation with inflammatory activity, serum ADA level may be a potential biomarker for predicting or monitoring histological activity in patients with AIH.展开更多
AIM： To investigate the usefulness of tumor markers and adenosine deaminase in differentiating between tuberculous peritonitis （TBP） and peritoneal carcinoma- tosis （PC）. METHODS： A retrospective analysis of dat...AIM： To investigate the usefulness of tumor markers and adenosine deaminase in differentiating between tuberculous peritonitis （TBP） and peritoneal carcinoma- tosis （PC）. METHODS： A retrospective analysis of data was per- formed on consecutive patients who underwent perito- neoscopic and abdominal computed tomography （CT） evaluations. Among 75 patients at the Seoul National University Hospital from January 2000 to June 2010 who underwent both tests, 27 patients （36.0%） and 25 patients （33.3%） were diagnosed with TBP and PC, re- spectively. Diagnosis was confirmed by peritoneoscopic biopsy. RESULTS： Serum c-reactive protein （7.88：1：6.62 mg/ dL vs 3.12 ＋ 2.69 mg/dL, P = 0.01）, ascites adenos- ine deaminase （66.76：1：32.09 IU/L vs 13.89 ：l： 8.95 IU/L, P 〈 0.01）, ascites lymphocyte proportion （67.77 ：1： 23.41% vs 48.36 ＋ 18.78%, P 〈 0.01）, and serum- ascites albumin gradient （0.72 ＋ 0.49 g/dL vs 1.05 ＋ 0.50 g/dL, P = 0.03） were significantly different be- tween the two groups. Among tumor markers, serum and ascites carcinoembryonic antigen, serum carbohy- drate antigen 19-9 showed significant difference be- tween two groups. Abdominal CT examinations showed that smooth involvement of the parietal peritoneum was more common in the TBP group （77.8% vs 40.7%） whereas nodular involvement was more common in the PC group （14.8% vs 40.7%, P = 0.04）. From receiver operating characteristic （ROC） curves ascites adeno- sines deaminase （ADA） showed better discriminative capability than tumor markers. An ADA cut-off level of 21 IU/L was found to yield the best results of differ- ential diagnosis; sensitivity, specificity, positive predic- tive value, and negative predictive value were 92.0%, 85.0%, 88.5% and 89.5%, respectively. CONCLUSION： Besides clinical and radiologic findings, ascitic fluid ADA measurement is helpful in the differen- tial diagnosis of TBP and PC.展开更多
Objective:To assess the efficacy of ADA isoenzyme estimation over that of total ADA level in pleural fluid and serum as a more efficient diagnostic indicator in tuberculous pleural effusions in high prevalence country...Objective:To assess the efficacy of ADA isoenzyme estimation over that of total ADA level in pleural fluid and serum as a more efficient diagnostic indicator in tuberculous pleural effusions in high prevalence country like India.Methods:The efficacy was analysed in total thirty four patients of pleural effusions.Total ADA was estimated by Guitsi and Galanti Calorimetric method and ADA isoenzymes with and without EHNA[Erythro-9-（2- hydroxy-3-nonyl） adenine]a potent ADA<sub>1</sub> inhibitor using the same method.Results:The results demonstrated a statistically significant values of ADA<sub>2</sub> in serum（P【0.001）,pleural fluid（P = 0.000） and significant value for the ratio of pleural fluid ADA<sub>2</sub>/serum ADA2（P【0.001） and pleural fluid ADA/ADA(<sub>2</sub>（P【0. 005）.The sensitivity and specificity values of pleural fluid ADA|2 is 81.8%and 91.6%（cut off value 60 IU/L for Tuberculous effusions）,serum ADA<sub>2</sub> 95.4%and 66%（cut off value 70 IU/L for tuberculous effusions）. ADA2<sub> </sub>is an isoenzyme,which is significantly raised in tuberculous pleural effusions both in the serum and pleural fluid.Conclusion:Estimation of ADA isoenzymes is redundant as a diagnostic aid over total ADA estimation in view of the limited improvements both in specificity and sensitivity patterns and also in term of cost-benefit ratio.展开更多
Objective:Relevance of estimation of pleural adenosine deaminase(PADA) and serum adenosine deaminase (SADA) levels in-pleural effusion especially in cases of lymphocytic predominant exudative tubercular effusions. Met...Objective:Relevance of estimation of pleural adenosine deaminase(PADA) and serum adenosine deaminase (SADA) levels in-pleural effusion especially in cases of lymphocytic predominant exudative tubercular effusions. Methods:Fifty patients(33 male and 17 female;age:44.12±11.51 years) with pleural effusions were selected to assay adenosine deaminase(ADA) activity in pleural fluid and serum in adjunct to pleural fluid analysis.Effusions were individually classified as transudates or exudates after careful evaluation of all the biochemical parameters of pleural fluid and serum of patients and on the basis of Lights criteria.Cutoff value for PADA was taken as 60U/L and that for pleural/serum ADA ratio(P/S ADA) was 1.8.Results:Fourty -three patients had exudative effusions among which 38 patients had tuberculous pleural effusions and 5 had nontubercular effusions.7 cases were transudates.Mean PADA levels in tubercular group(78.95±25.32 U/ L) were found to be much higher P=0.0000) than nontubercular(23.00±5.22 U/L) group.SADA levels in tubercular group(31.05±6.42 U/L) were significantly higher(P=0.0000)as compared to nontubercular group(15.58±8.35 U/L).PADA cutoff at 60 U/L yielded sensitivity and.specificity of 81.5%and 100%respectively,whereas P/S ADA ratio at 1.8 gave sensitivity and specificity of 84.2% and 75%respectively. A positive correlation(r=0.507,P= 0.001 1)between PADA and SADA was found in tubercular group but no such correlation(r=0.302,P=0.3407)was observed in nontubercular group.Conclusion: The measurement of ADA in tubercular pleural effusions has not only relevance but also a high diagnostic utility when other clinical and laboratory tests are either negative or confusing.展开更多
Interactions of anionic, cationic and metal phthalocyanine with adenosine deaminase were studied by molecular dynamics and docking simulation. Structural parameters such as solvent accessible surface area (SAS), mid-p...Interactions of anionic, cationic and metal phthalocyanine with adenosine deaminase were studied by molecular dynamics and docking simulation. Structural parameters such as solvent accessible surface area (SAS), mid-point of transition temperature (Tm), radial distribution function (RDF) and hydrogen bond, helix, coil, beta percentage and other physical parameters were obtained. The denaturation of adenosine deaminase (ADA) by heat, anionic and cationic phthalocyanines was compared. A series of 20 ns simulation performed at temperatures ranging from 275 to 450 K, starting from the ADA native structure. Results of radial distribution functions (RDFs) showed that metallic derivative at low concentration behaves the same as osmolytes that increases the beta form and increases the enzyme stability. Molecular docking studies have been carried out to confirm the simulation results. Investigation of binding site and free energy confirmed that the efficiency of interaction with adenosine deaminase depends on metal core. Binding energy of non-metallic form is more negative than metallic form and it significantly decreases for phthalocyanine. Self-aggregation of anionic phthalocyanine decreases in comparison with cationic derivative, therefore enzyme denaturation in the presence of anionic form is higher than the other. Furthermore, thermal stability of the enzyme also depends on temperature in presence of phthalocyanine. Binding site of phthalocyanine on the enzyme has been identified by docking analysis.展开更多
Background: Adenosine deaminase (ADA) and 5'-nucleotidase (5'-NT) play a crucial role in adenosine metabolism in healthy individuals. Adenosine is an inflammatory mediator of asthma. Changes in adenosine metab...Background: Adenosine deaminase (ADA) and 5'-nucleotidase (5'-NT) play a crucial role in adenosine metabolism in healthy individuals. Adenosine is an inflammatory mediator of asthma. Changes in adenosine metabolism and role of ADA and 5'-NT in regulating adenosine level in asthmatics and correlation of these changes with severity of asthma are not clearly understood. Methods: In this study, we screened 5217 patients, of which 2416 were diagnosed with asthma. Further, of 2416 asthmatics, only 45 patients who strictly fulfilled the selection criteria were enrolled in the study. The patients were classified into mild, moderate and severe persistent groups;each group consisted of fifteen patients. Fifteen healthy subjects served as controls. Adenosine levels and activities of 5'-NT, total ADA, ADA1 and ADA2 in serum, lymphocytes and erythrocytes were determined. The data were analysed statistically and p vice versa.展开更多
目的:探究血清ADA、LDH及EBV-DNA水平对传染性单核细胞增多症肝损害患儿病情及预后评估的临床价值。方法:选取109例传染性单核细胞增多症(IM)患儿为研究对象,按是否出现肝损害将患儿分为肝损害组(n=62)与非肝损害组(n=47)。检测两组患...目的:探究血清ADA、LDH及EBV-DNA水平对传染性单核细胞增多症肝损害患儿病情及预后评估的临床价值。方法:选取109例传染性单核细胞增多症(IM)患儿为研究对象,按是否出现肝损害将患儿分为肝损害组(n=62)与非肝损害组(n=47)。检测两组患儿血清腺苷脱氨酶(ADA)、乳酸脱氢酶(LDH)水平及EB病毒脱氧核糖核酸(EBV-DNA)载量,分析不同肝损害程度上述指标表达水平及其与传染性单核细胞增多症并发肝损害的相关性,绘制ROC曲线分析上述指标的诊断效能。结果:肝损害组患儿发生肝肿大率、ALT、AST、TBil水平均高于非肝损害组(P<0.05);肝损害组患儿血清ADA、LDH及EBV-DNA水平高于非肝损害组(P<0.05),且均随肝损害程度升高而升高(P<0.05);传染性单核细胞增多症患儿血清ADA、LDH及EBV-DNA水平与ALT、AST均呈正相关(P<0.05);ROC曲线显示,血清ADA、LDH及EBV-DNA的曲线下面积(AUC)分别为0.831、0.861和0.934,截断值分别为30.60 U/L、544.63 U/L和2.35 lg copies/mL,联合诊断的AUC为0.957。结论:血清ADA、LDH及EBV-DNA水平与IM患儿肝损害相关,对于评估IM合并肝损害诊断以及预后评估方面具有一定参考价值,值得临床广泛应用。展开更多
既往研究发现,SMAD特异性E3泛素蛋白连接酶1(SMAD specific E3 ubiquitin protein ligase 1,SMURF1)通过其E3泛素连接酶活性介导自噬进程,然而SMURF1的泛素化底物蛋白质仍有待进一步挖掘。本文利用免疫共沉淀(Co-IP)联合蛋白质谱分析捕...既往研究发现,SMAD特异性E3泛素蛋白连接酶1(SMAD specific E3 ubiquitin protein ligase 1,SMURF1)通过其E3泛素连接酶活性介导自噬进程,然而SMURF1的泛素化底物蛋白质仍有待进一步挖掘。本文利用免疫共沉淀(Co-IP)联合蛋白质谱分析捕获并鉴定THP-1细胞中SMURF1的相互作用蛋白质集合物,发现在THP-1细胞中SMURF1可与222种蛋白质物理性结合,RNA腺苷脱氨酶1(adenosine deaminase acting on RNA 1,ADAR1)具有较高的肽段结合分数。构建SMURF1过表达载体并转染到HEK-293T细胞中,Co-IP和Western印迹检测验证外源性SMURF1与内源性ADAR1存在相互作用。qRT-PCR和Western印迹检测结果显示,在HEK-293T细胞中过表达SMURF1后ADAR 1 mRNA水平差异无统计学意义、蛋白质水平明显降低(P<0.05)。用放线菌酮(CHX)分别处理正常和过表达SMURF1的HEK-293T细胞,Western印迹检测显示,过表达SMURF1后ADAR1的半衰期缩短(P<0.05)。进一步在HEK-293T细胞中共转染泛素(Ub)过表达载体和SMURF1过表达载体,通过Co-IP和Western印迹检测结果证实,过表达SMURF1后ADAR1的多聚泛素化水平显著增加(P<0.05)。在蛋白酶体抑制剂(MG132)作用后,Western印迹检测结果表明,蛋白酶体降解途径受抑制后SMURF1对ADAR1的负调控作用减弱(P<0.05)。本研究表明,SMURF1可以与ADAR1相互作用,催化ADAR1的多聚泛素化修饰并介导其蛋白酶体途径降解,为探索SMURF1通过影响ADAR1蛋白质稳定性而具备的多种生物学功能提供理论基础。展开更多
文摘AIM To evaluate the relationship between serum adenosine deaminase(ADA) levels and histological features in patients with autoimmune hepatitis(AIH).METHODS A total of 80 subjects(52 AIH cases and 28 healthy controls) were included in the study. Patients were diagnosed according to the simplified criteria suggested by the International Autoimmune Hepatitis Group. All of the cases had been diagnosed with AIH between 2010-2015 at Hacettepe University, Department of Gastroenterology. Serum blood samples were collected and stored at-80 ℃ until the biochemical estimation of ADA activity. The diagnosis of patients was confirmed by liver biopsy. Serum ADA > 20 U/L was considered to be high level.RESULTS Mean serum ADA levels were significantly higher in AIH patients than those in healthy controls(25.4 ± 9.6 U/L vs 12.8 ± 2.2 U/L, P < 0.001). Serum ADA levels > 20 U/L were found in 63.5% AIH patients and in 0% healthy controls(P < 0.001). Mean serum ADA levels were significantly increased in each stage of histological activity: 15.2 ± 3.5 U/L for patients with mild interface hepatitis, 23.1 ± 10.0 U/L for patients with moderate interface hepatitis and 30.9 ± 7.0 U/L for patients with severe interface hepatitis(P < 0.001). Correlation analysis showed that there was a positive association between serum ADA levels and histological activity(r = 0.71, P < 0.001). Receiver operating characteristic analysis suggested that 24.5 U/L was the optimum cut-off point of ADA level for severe interface hepatitis(sensitivity 88%, specificity 85.2%, area under thecurve: 0.88).CONCLUSION Because of the positive correlation with inflammatory activity, serum ADA level may be a potential biomarker for predicting or monitoring histological activity in patients with AIH.
文摘AIM： To investigate the usefulness of tumor markers and adenosine deaminase in differentiating between tuberculous peritonitis （TBP） and peritoneal carcinoma- tosis （PC）. METHODS： A retrospective analysis of data was per- formed on consecutive patients who underwent perito- neoscopic and abdominal computed tomography （CT） evaluations. Among 75 patients at the Seoul National University Hospital from January 2000 to June 2010 who underwent both tests, 27 patients （36.0%） and 25 patients （33.3%） were diagnosed with TBP and PC, re- spectively. Diagnosis was confirmed by peritoneoscopic biopsy. RESULTS： Serum c-reactive protein （7.88：1：6.62 mg/ dL vs 3.12 ＋ 2.69 mg/dL, P = 0.01）, ascites adenos- ine deaminase （66.76：1：32.09 IU/L vs 13.89 ：l： 8.95 IU/L, P 〈 0.01）, ascites lymphocyte proportion （67.77 ：1： 23.41% vs 48.36 ＋ 18.78%, P 〈 0.01）, and serum- ascites albumin gradient （0.72 ＋ 0.49 g/dL vs 1.05 ＋ 0.50 g/dL, P = 0.03） were significantly different be- tween the two groups. Among tumor markers, serum and ascites carcinoembryonic antigen, serum carbohy- drate antigen 19-9 showed significant difference be- tween two groups. Abdominal CT examinations showed that smooth involvement of the parietal peritoneum was more common in the TBP group （77.8% vs 40.7%） whereas nodular involvement was more common in the PC group （14.8% vs 40.7%, P = 0.04）. From receiver operating characteristic （ROC） curves ascites adeno- sines deaminase （ADA） showed better discriminative capability than tumor markers. An ADA cut-off level of 21 IU/L was found to yield the best results of differ- ential diagnosis; sensitivity, specificity, positive predic- tive value, and negative predictive value were 92.0%, 85.0%, 88.5% and 89.5%, respectively. CONCLUSION： Besides clinical and radiologic findings, ascitic fluid ADA measurement is helpful in the differen- tial diagnosis of TBP and PC.
文摘Objective:To assess the efficacy of ADA isoenzyme estimation over that of total ADA level in pleural fluid and serum as a more efficient diagnostic indicator in tuberculous pleural effusions in high prevalence country like India.Methods:The efficacy was analysed in total thirty four patients of pleural effusions.Total ADA was estimated by Guitsi and Galanti Calorimetric method and ADA isoenzymes with and without EHNA[Erythro-9-（2- hydroxy-3-nonyl） adenine]a potent ADA<sub>1</sub> inhibitor using the same method.Results:The results demonstrated a statistically significant values of ADA<sub>2</sub> in serum（P【0.001）,pleural fluid（P = 0.000） and significant value for the ratio of pleural fluid ADA<sub>2</sub>/serum ADA2（P【0.001） and pleural fluid ADA/ADA(<sub>2</sub>（P【0. 005）.The sensitivity and specificity values of pleural fluid ADA|2 is 81.8%and 91.6%（cut off value 60 IU/L for Tuberculous effusions）,serum ADA<sub>2</sub> 95.4%and 66%（cut off value 70 IU/L for tuberculous effusions）. ADA2<sub> </sub>is an isoenzyme,which is significantly raised in tuberculous pleural effusions both in the serum and pleural fluid.Conclusion:Estimation of ADA isoenzymes is redundant as a diagnostic aid over total ADA estimation in view of the limited improvements both in specificity and sensitivity patterns and also in term of cost-benefit ratio.
文摘Objective:Relevance of estimation of pleural adenosine deaminase(PADA) and serum adenosine deaminase (SADA) levels in-pleural effusion especially in cases of lymphocytic predominant exudative tubercular effusions. Methods:Fifty patients(33 male and 17 female;age:44.12±11.51 years) with pleural effusions were selected to assay adenosine deaminase(ADA) activity in pleural fluid and serum in adjunct to pleural fluid analysis.Effusions were individually classified as transudates or exudates after careful evaluation of all the biochemical parameters of pleural fluid and serum of patients and on the basis of Lights criteria.Cutoff value for PADA was taken as 60U/L and that for pleural/serum ADA ratio(P/S ADA) was 1.8.Results:Fourty -three patients had exudative effusions among which 38 patients had tuberculous pleural effusions and 5 had nontubercular effusions.7 cases were transudates.Mean PADA levels in tubercular group(78.95±25.32 U/ L) were found to be much higher P=0.0000) than nontubercular(23.00±5.22 U/L) group.SADA levels in tubercular group(31.05±6.42 U/L) were significantly higher(P=0.0000)as compared to nontubercular group(15.58±8.35 U/L).PADA cutoff at 60 U/L yielded sensitivity and.specificity of 81.5%and 100%respectively,whereas P/S ADA ratio at 1.8 gave sensitivity and specificity of 84.2% and 75%respectively. A positive correlation(r=0.507,P= 0.001 1)between PADA and SADA was found in tubercular group but no such correlation(r=0.302,P=0.3407)was observed in nontubercular group.Conclusion: The measurement of ADA in tubercular pleural effusions has not only relevance but also a high diagnostic utility when other clinical and laboratory tests are either negative or confusing.
文摘Interactions of anionic, cationic and metal phthalocyanine with adenosine deaminase were studied by molecular dynamics and docking simulation. Structural parameters such as solvent accessible surface area (SAS), mid-point of transition temperature (Tm), radial distribution function (RDF) and hydrogen bond, helix, coil, beta percentage and other physical parameters were obtained. The denaturation of adenosine deaminase (ADA) by heat, anionic and cationic phthalocyanines was compared. A series of 20 ns simulation performed at temperatures ranging from 275 to 450 K, starting from the ADA native structure. Results of radial distribution functions (RDFs) showed that metallic derivative at low concentration behaves the same as osmolytes that increases the beta form and increases the enzyme stability. Molecular docking studies have been carried out to confirm the simulation results. Investigation of binding site and free energy confirmed that the efficiency of interaction with adenosine deaminase depends on metal core. Binding energy of non-metallic form is more negative than metallic form and it significantly decreases for phthalocyanine. Self-aggregation of anionic phthalocyanine decreases in comparison with cationic derivative, therefore enzyme denaturation in the presence of anionic form is higher than the other. Furthermore, thermal stability of the enzyme also depends on temperature in presence of phthalocyanine. Binding site of phthalocyanine on the enzyme has been identified by docking analysis.
文摘Background: Adenosine deaminase (ADA) and 5'-nucleotidase (5'-NT) play a crucial role in adenosine metabolism in healthy individuals. Adenosine is an inflammatory mediator of asthma. Changes in adenosine metabolism and role of ADA and 5'-NT in regulating adenosine level in asthmatics and correlation of these changes with severity of asthma are not clearly understood. Methods: In this study, we screened 5217 patients, of which 2416 were diagnosed with asthma. Further, of 2416 asthmatics, only 45 patients who strictly fulfilled the selection criteria were enrolled in the study. The patients were classified into mild, moderate and severe persistent groups;each group consisted of fifteen patients. Fifteen healthy subjects served as controls. Adenosine levels and activities of 5'-NT, total ADA, ADA1 and ADA2 in serum, lymphocytes and erythrocytes were determined. The data were analysed statistically and p vice versa.
文摘目的:探究血清ADA、LDH及EBV-DNA水平对传染性单核细胞增多症肝损害患儿病情及预后评估的临床价值。方法:选取109例传染性单核细胞增多症(IM)患儿为研究对象,按是否出现肝损害将患儿分为肝损害组(n=62)与非肝损害组(n=47)。检测两组患儿血清腺苷脱氨酶(ADA)、乳酸脱氢酶(LDH)水平及EB病毒脱氧核糖核酸(EBV-DNA)载量,分析不同肝损害程度上述指标表达水平及其与传染性单核细胞增多症并发肝损害的相关性,绘制ROC曲线分析上述指标的诊断效能。结果:肝损害组患儿发生肝肿大率、ALT、AST、TBil水平均高于非肝损害组(P<0.05);肝损害组患儿血清ADA、LDH及EBV-DNA水平高于非肝损害组(P<0.05),且均随肝损害程度升高而升高(P<0.05);传染性单核细胞增多症患儿血清ADA、LDH及EBV-DNA水平与ALT、AST均呈正相关(P<0.05);ROC曲线显示,血清ADA、LDH及EBV-DNA的曲线下面积(AUC)分别为0.831、0.861和0.934,截断值分别为30.60 U/L、544.63 U/L和2.35 lg copies/mL,联合诊断的AUC为0.957。结论:血清ADA、LDH及EBV-DNA水平与IM患儿肝损害相关,对于评估IM合并肝损害诊断以及预后评估方面具有一定参考价值,值得临床广泛应用。
文摘既往研究发现,SMAD特异性E3泛素蛋白连接酶1(SMAD specific E3 ubiquitin protein ligase 1,SMURF1)通过其E3泛素连接酶活性介导自噬进程,然而SMURF1的泛素化底物蛋白质仍有待进一步挖掘。本文利用免疫共沉淀(Co-IP)联合蛋白质谱分析捕获并鉴定THP-1细胞中SMURF1的相互作用蛋白质集合物,发现在THP-1细胞中SMURF1可与222种蛋白质物理性结合,RNA腺苷脱氨酶1(adenosine deaminase acting on RNA 1,ADAR1)具有较高的肽段结合分数。构建SMURF1过表达载体并转染到HEK-293T细胞中,Co-IP和Western印迹检测验证外源性SMURF1与内源性ADAR1存在相互作用。qRT-PCR和Western印迹检测结果显示,在HEK-293T细胞中过表达SMURF1后ADAR 1 mRNA水平差异无统计学意义、蛋白质水平明显降低(P<0.05)。用放线菌酮(CHX)分别处理正常和过表达SMURF1的HEK-293T细胞,Western印迹检测显示,过表达SMURF1后ADAR1的半衰期缩短(P<0.05)。进一步在HEK-293T细胞中共转染泛素(Ub)过表达载体和SMURF1过表达载体,通过Co-IP和Western印迹检测结果证实,过表达SMURF1后ADAR1的多聚泛素化水平显著增加(P<0.05)。在蛋白酶体抑制剂(MG132)作用后,Western印迹检测结果表明,蛋白酶体降解途径受抑制后SMURF1对ADAR1的负调控作用减弱(P<0.05)。本研究表明,SMURF1可以与ADAR1相互作用,催化ADAR1的多聚泛素化修饰并介导其蛋白酶体途径降解,为探索SMURF1通过影响ADAR1蛋白质稳定性而具备的多种生物学功能提供理论基础。
