首页 | 本学科首页   官方微博 | 高级检索  
相似文献
 共查询到20条相似文献,搜索用时 234 毫秒
1.
高分子量和低分子量尿激酶的分离纯化及动力学性质研究   总被引:1,自引:0,他引:1  
人尿激酶粗品经苯甲脒亲和柱纯化和Protein-PakSP柱分离后,得到两种分子量的尿激酶(UK),即高分子量尿激酶(HUK)和低分子量尿激酶(LUK),采用民斯亮蓝法测定蛋白质浓度,纤维蛋白板法测定活力,测得HUK比活为2.9×10^5IU/mg蛋白,LUK为3.510^5IU/mg蛋白,活力回收为70%以上,经SDS-PAGE鉴定,HUK和LUK均呈单一条带,分子量分别为54kD和33kD,H  相似文献   

2.
何首乌(PolygonummultiflorumThunb.)叶Cu·ZnSOD经硫酸铵盐析、离子交换柱层析及葡聚糖凝胶过滤等步骤,被分离成两组SOD同工酶(SOD1,SOD2),它们被纯化到均一程度。SOD1分子量为32.4kD,亚基分子量为16.2kD,最适pH值为50,在60℃和65℃时的半衰期分别为14min和8min;SOD2分子量为305kD,亚基分子量为159kD,最适pH值为60,在60℃和65℃时半衰期分别为32min和16min,它由274个氨基酸残基组成,不含Cys、Tyr和Try。SOD1和SOD2每mol酶都含有2molCu和2molZn,它们在紫外区最大吸收波长均为275nm。  相似文献   

3.
低分子量硫酸葡聚糖对小鼠造血干细胞动员作用的研究   总被引:3,自引:0,他引:3  
给小鼠静脉注射低分子量(<10 ̄4u)硫酸葡聚糖(DS)15mg/kg后外周血中白细胞、单个核细胞(mononuclearcek,MNC)、CFU-GM、BFU-E和CFU-Mix产率等指标出现时相性变化。给药后1h开始升高,2h达到高峰、分别为药前值的2.2、2.6、3.8、4.4和3.0倍,7h时趋向正常。给药后2h上述各类细胞在外周血中的含量随着DS的剂量增加而增加。白细胞、MNC计数在DS180mg/kg时达到峰值,均为对照组的4倍。240mg/k8时未见明显增加。不同剂量DS对各系祖细胞均有不同程度的动员作用,DS剂量15-30mg/kg效果最好,每升血中CFU-GM、BFU-E、CFU-Mix的数量分别相当于对照组的5.0、11.9和8.8倍。其峰值出现时间与白细胞、MNC不同,表明DS对不同类型细胞的作用机制也不尽一致。经口给小鼠投以DS240和48omg/kg后,未见外周血中白细胞、MNC计数有显著性升高,提示对造血干细胞没有动员作用。  相似文献   

4.
以魔芋葡甘聚糖(KGM)凝胶作为铜金属螯合亲和层析的载体一步亲和纯化猪血SOD,得到电泳均一,比活为8622U/mg,纯化倍数为77.8倍的SOD,其回收率为85.4%.探讨了魔芋葡甘聚糖凝胶作为亲和载体的可能性及前景.  相似文献   

5.
猴头子实体锰型超氧物歧化酶的纯化及其性质鉴定   总被引:6,自引:0,他引:6  
猴头子实体的超氧物歧化酶仅Mn-SOD1种,有其资源和学术研究上的意义。其粗酶液经(NH4)2SO4盐析,DE52和CM52离子交换柱层析,纯化到电泳单斑点均一程度,其比活性为3096.5u/mg,活性回收率为14.8%。纯化的Mn-SOD分子量为47.0KD,亚基分子量为23.3KD。金属元素分析表明每个亚基含1个Mn原子。该酶在紫外区有2个吸收峰,分别在218.4nm和276.0nm。该酶的理  相似文献   

6.
以魔芋葡甘聚糖凝胶为载体亲和层析分离纯化猪血SOD   总被引:4,自引:0,他引:4  
以魔芋葡甘聚糖(KGM)凝胶作为铜金属螯合亲和层析的载体一步亲和纯化猪血SOD,得到电泳均一,比活为8622U/mg,纯化倍数为77.8倍的SOD,其回收率为85.4%。探讨了魔芋葡甘聚糖凝胶作为亲和载体的可能性及前景。  相似文献   

7.
韭菜线粒体锰超氧化物歧化酶纯化及性质研究   总被引:4,自引:0,他引:4  
经硫酸铵沉淀、DEAE-Sephacel层析和Sephadex G-200凝胶过滤,将韭菜线粒体SOD纯化到均一程度。从6000g韭菜叶片线粒体中纯化得到2.5mg酶,酶比活力达1200U/mg蛋白。该酶对KCN和H2O2都不敏感,热稳定性弱 外光区吸收峰在280nm,凝胶过滤法测得其分子量为8200D,SOS-PAGE法测定其亚基分子量的22000D,DNS法测得其N-末端氨基酸为缬氨酸。上述结  相似文献   

8.
用国产微孔玻璃珠从CD-1工程细胞株培养上清中纯化u-PA,摸索了微孔玻璃珠结合u-PA的条件和洗脱方法,比较了硫酸铵线性梯度洗脱或25%乙二醇洗脱u-PA活性的结果,最后确定洗脱的条件为0.25mol/LTris、1~2mol/L(NH4)2SO4、pH9.3。经此一步纯化,u-PA比活性达到15329IU/mg,回收率78%,还原条件下SDS-PAGE分子量为52×103的单链u-PA占74%。  相似文献   

9.
绿豆苯丙氨酸解氨酶的性质及抗肿瘤作用研究   总被引:10,自引:0,他引:10  
本文参照Havir的方法,从绿豆中分离纯化了苯丙氨酸解氨酶(PAL)。纯化的PAL经SDS聚丙烯酰胺凝胶电泳及等电聚焦电泳鉴定为单一的蛋白质区带,并测得亚基分子量为76KD,等电点为5.45。在PAL对L1210小鼠淋巴细胞白血病细胞株的体外抑制实验观察到:PAL对该瘤株的抑制作用随作用时间的延长和药物剂量的增加而增强,0.2U/ml、1.0U/ml、2.0U/ml、4.0U/ml、6.0U/ml、10.0U/ml的PAL作用癌细胞72h,其抑制率分别为25.8%、40.0%、55.3%、72.6%、77.9%、82.9%。  相似文献   

10.
用硫酸铵分级沉淀、DEAE-纤维素离子交换层析、免疫亲和层析、SephadexG100凝胶柱层析从人胃组织中提取出腺苷脱氨酶,酶纯化19324倍,比活力为5797U/mg蛋白.提取酶液经PAGE、SDS-PAGE和等电聚焦只呈现一条区带。测得该酶的分子量为41.2kD,等电点为pH4.8.氨基酸组成分析表明该酶由388个氨基酸残基组成,N端氨基酸为精氨酸。酶的最适pH为6.5,pH小于5.0或大于9.0时不稳定;最适温度为37℃,对热不太稳定,以腺苷及2-脱氧腺苷作为底物,其Km分别为87μmol/L和41μmol/L。  相似文献   

11.
肝再生增强因子(ALR)是一类胞源性肝细胞生长因子。为在毕赤酵母中分泌表达人肝再生增强因子(rhALR),以色谱法分离纯化后进行体外活性研究,构建表达载体pPICZαA- ALR,经电穿孔转入毕赤酵母中,用0.5%甲醇诱导表达;重组酵母培养上清经SDS-PAGE电泳和western blot鉴定后表明, rhALR以分子量为30kD的二聚体为主;定量分析结果表明,重组酵母培养上清中rhALR约占总蛋白的66%,表达量约为40mg/L;经DEAE柱和G75柱纯化后,获得的rhALR纯度大于95%,得率为52%;体外生物学活性实验表明,rhALR能明显促进HepG2、SMMC-7721和NIH-3T3细胞的增殖。  相似文献   

12.
油柑果超氧化物歧化酶的化学修饰及其性质的研究   总被引:2,自引:0,他引:2  
从油柑果中提取SOD[Superoxide dismutase(EC1,15,1,1,)],用山梨醇月桂酸酯(Sorbitol laurate)进行化学修饰,得到SL-SOD。比活力为4200u/mg。交联葡聚糖凝胶层析测分子量为38KD,紫外区最大吸收峰是258nm,修饰后SOD对温度和PH的稳定性均有增强。在某些低浓度的有机介质中活性比在水中高。修饰SOD的半衰期为14h,有效保存期为43d。几乎无免疫原性。  相似文献   

13.
Guo FX  Shi-Jin E  Liu SA  Chen J  Li DC 《Mycologia》2008,100(3):375-380
A thermostable superoxide dismutase (SOD) from the culture supernatant of a thermophilic fungus Chaetomium thermophilum strain CT2 was purified to homogeneity by fractional ammonium sulfate precipitation, ion-exchange chromatography on DEAE-sepharose, phenyl-sepharose hydrophobic interaction chromatography. The pure SOD had a specific activity of 115.77 U/mg of protein and was purified 7.49-fold, with a yield of 14.4%. The molecular mass of a single band of the enzyme was estimated to be 23.5 kDa, using sodium dodecyl sulfate-polyacrylamide gel electrophoresis. Using gel filtration on Sephacryl S-100, the molecular mass was estimated to be 94.4 kDa, indicating that this enzyme was composed of four identical subunits of 23.5 kDa each. The SOD was found to be inhibited by NaN3, but not by KCN and H2O2. Atomic absorption spectrophotometric analysis showed that the content of Mn was 2.05 microg/mg of protein and Fe was not detected in the purified enzyme. These results suggested that the SOD in C. thermophilum was the manganese superoxide dismutase type. N-terminal amino acid sequencing (10 residues) was KX (X is uncertain) TLPDLKYD. The N-terminal amino acid sequencing homologies to other MnSod also indicated that it was a manganese-containing superoxide dismutase. The SOD exhibited maximal activity at pH 7.5 and optimum temperature at 60 C. It was thermostable at 50 and 60 C and retained 60% activity after 60 min at 70 C. The half-life of the SOD at 80 C was approximately 25 min and even retained 20% activity after 30 min at 90 C.  相似文献   

14.
Superoxide dismutase (SOD) was purified from cells of the strict anaerobic methanogenic archaeon Methanobrevibacter arboriphilus strain AZ. The four-step purification procedure resulted in enzyme with specific activity of 3970 units/mg and yield of 22%. It was shown that the SOD is a Fe-containing homotetramer composed of subunits of 21.2 kD each. Sodium azide (13.5 mM), unlike KCN, inhibits the activity of the SOD. Hydrogen peroxide (0.5 mM) inactivates the enzyme, which is consistent with the properties of the known Fe-containing SODs from methanogenic Archaea.  相似文献   

15.
This study examined the effects of lung collapse, a condition that causes relative hypoxia in lung tissues, on superoxide dismutase (SOD), cytochrome oxidase (cyt ox), and pyruvate kinase (py ki) activities in rabbits. Cyanide-insensitive respiration measurements were done in collapsed and contralateral lungs, as an index of intracellular free radical production. Rabbits' right lungs were collapsed for 7 days after which the animals were killed. We found that control rabbit lungs contained approximately 25 SOD units/mg DNA measured with 10(-5) M KCN (total SOD) and approximately 11 SOD units/mg DNA measured with 10(-3) M KCN (mitochondrial or MnSOD). Right lung collapse caused a 25% decrease in mitochondrial SOD activity after 7 days (P less than 0.05), whereas no significant changes occurred in right or left lungs' total SOD activity. In control rabbits cyt ox activity averaged approximately 0.009 mumol ferrocytochrome c.min-1.mg DNA-1. Right lung collapse caused a greater than 40% decrease in cyt ox activity after 7 days of collapse (P less than 0.05), whereas cyt ox activity in contralateral left lungs did not change. Pyruvate kinase activity, a marker for anaerobic glycolysis resulting from tissue hypoxia, increased 49% in collapsed right lungs (P less than 0.01). Cyanide-insensitive respiration was 83% higher in 7 day-collapsed lungs (2.28 +/- 0.66 microliters O2.min-1.g-1) compared with contralateral lungs (1.24 +/- 0.34, P less than 0.05), indicating increased O2-. and H2O2 production in this tissue after homogenization at normoxic PO2 (approximately 150 Torr).(ABSTRACT TRUNCATED AT 250 WORDS)  相似文献   

16.
目的探讨硫化氢(H2S)对阿霉素(DOX)诱导的H9c2细胞损伤的影响及其作用机制。 方法H2S对DOX心肌毒性保护作用的实验分组为:对照组(Control组),5?μmol/?L DOX处理组(A组),5?μmol/L DOX和400?μmol/L NaHS共同处理组(B组),400?μmol/L NaHS单独处理组(C组),5?μmol/L DOX、400?μmol/L NaHS和15?μmol/L Sirtinol共同处理组(D组),15?μmol/L Sirtinol单独处理组(E组)。SIRT1是否参与H2S抗DOX心肌毒性作用机制的实验分组为:对照组(Control组),5?μmol/L DOX处理组(F组),5?μmol/L DOX和400?μmol/L NaHS共同处理组(G组),5?μmol/L DOX、400?μmol/L NaHS和15?μmol/L Sirtinol共同处理组(H组),15?μmol/L Sirtinol单独处理组(I组)。使用MTT法检测细胞活力;Elisa法检测细胞MDA以及SOD水平;DCFH-?DA荧光探针法检测ROS水平;采用Western Blot法检测SIRT1蛋白表达。使用单因素方差分析法进行统计学分析。 结果NaHS预处理可抑制DOX导致的H9c2细胞活力下降:Control组,A组、B组、C组细胞活力分别为100﹪、(54.58±1.58)﹪、(85.05±4.31)﹪、(100.22±4.46)﹪ (F = 134.9,P < 0.001)。NaHS预处理可减弱DOX引起的H9c2细胞ROS、MDA水平的增加以及SOD水平的降低:Control组的ROS、MDA和SOD水平分别是100﹪、(34.18±1.56) μmol/g、(53.69±1.44) U/?mg;A组的ROS、MDA和SOD水平分别是(174.90±12.65)﹪、(72.65±2.66) μmol/g、(31.80±2.05) U/?mg;B组的ROS、MDA和SOD水平分别是(126.08±6.25)﹪、(44.59±1.92) μmol/g、(48.06±1.56) U/mg;C组的ROS、MDA和SOD水平分别是(91.86±1.66)﹪、(32.93±1.56)?μmol/?g、(55.93±1.58)?U/?mg (F?= 83.26,P < 0.001;F = 271.4,P < 0.001;F = 127.0,P < 0.001)。F组(6、12、24?h)H9c2细胞SIRT1蛋白表达水平分别是(0.45±0.03)、(0.27±0.02)、(0.25±0.03),较Control组(1.00±0.00)降低(F = 611.1,P < 0.001)。本研究还发现,NaHS预处理H9c2细胞能阻止DOX引起的SIRT1蛋白表达下调:Control组、F组、G组、H组的SIRT1蛋白表达水平分别是(1.00±0.00)、(0.31±0.03)、(0.60±0.04)、(1.09±0.09)(F = 123.4,P?2S对DOX诱导的H9c2细胞活力降低的抑制作用:Control组,F组、G组、H组、I组细胞活力分别为100﹪、(54.58±1.58)﹪、(85.37±3.62)﹪、(71.11±2.11)﹪、(97.53±1.45)﹪ (F = 238.2,P < 0.001)。Sirtinol预处理可明显逆转H2S对DOX导致的H9c2细胞ROS和MDA含量增加及SOD水平降低的抑制作用:Control组的ROS、MDA和SOD水平分别是100﹪、(35.84±2.22)μmol/?g、(53.03±3.16) U/mg;F组的ROS、MDA和SOD水平分别是(184.6±11.33)﹪、(74.78±5.30)μmol/g、(29.26±0.85)U/mg;G组的ROS、MDA和SOD水平分别是(126.5±7.57)﹪、(41.95±3.43)μmol/g、(52.61±2.26)U/mg;H组的ROS、MDA和SOD水平分别是(174.7±5.50)﹪、(67.69±1.52) μmol/g、(35.33±1.95) U/mg,I组的ROS、MDA和SOD水平分别是(98.03±2.86)﹪、(37.66±2.49)μmol/g、51.14 U/mg(F = 112.0,P < 0.001;F = 93.73,P < 0.001;F = 84.92,P < 0.001)。 结论H2S通过调控SIRT1抑制DOX诱导的H9c2细胞损伤。  相似文献   

17.
The hypothesis of the present study is that cardiomyocytes subjected to prolonged ischemia, may release survival factors that will protect new cardiac cells from ischemic stress. We exposed neonatal rat cardiomyocyte primary cultures to hypoxia, collected the supernatant, treated intact cardiac cells by this posthypoxic supernatant, and exposed them to hypoxia. The results show cardioprotection of the treated cells compared with the untreated ones. We named the collected posthypoxic supernatant "conditioned medium" (CM), which acts in a dose-dependent manner to protect new cardiac cells from hypoxia: 100 or 75% of CM diluted in phosphate-buffered saline (PBS) protected cells as if they were not exposed to hypoxia (P < 0.001). When CM was removed from the cells before hypoxia, protection was not observed. CM also protected skeletal muscle cultures from hypoxia, but not cardiac cells against H(2)O(2)-induced cell damage. Finally, CM treatment protected the isolated heart in Langendorff set-up against ischemia. Smaller infarct size (9.9 ± 4.4% vs. 28.3 ± 8.5%, P < 0.05), better Rate Pressure Product (67 ± 11% vs. 48.6 ± 13.4%, P < 0.05) and better rate of contraction and relaxation were observed following ischemia and reperfusion (1341 ± 399 mmHg/s vs. 951 ± 349 mmHg/s, P < 0.05 and 1053 ± 347 mmHg/s vs. 736 ± 314 mmHg/s, P < 0.05). To conclude, there are factors that are released from the heart cells subjected to ischemia/hypoxia that protects cardiomyocytes from ischemic stress.  相似文献   

18.
近江牡蛎铜锌超氧化物歧化酶的纯化及部分性质研究   总被引:7,自引:0,他引:7  
经65℃加热,硫酸铵分级沉淀,SephadexG-100凝胶过滤和DE-52柱层析,从近江牡蛎(OstrearivularisGould)软体部分提纯了铜锌超氧化物歧化酶(Cu,Zn-SOD).对其理化性质鉴定表明,用此法纯化的酶纯度均一.该酶系由两个相同亚基组成的二聚体,分子量27.9kD.该酶的紫外吸收峰在272.5nm,红外光谱表现出其氨基酸组成特征,与猪血SOD存在差异.该酶在不同的升温速率下及经不同浓度的H2O2处理后的稳定性与猪血SOD不同.其氨基酸组成与不同来源的同类酶存在差异.  相似文献   

19.
Sinha S 《Life sciences》2005,76(12):1407-1426
The expression of copper zinc superoxide dismutase (SOD1), manganese superoxide dismutase (SOD2), glutathione peroxidase (GPx), and catalase (CAT) genes have been detected in human skin fibroblast cells for 2 year normal child (control), 50 year old normal male and female and a 1 year old Down Syndrome (DS) male and female with established trisomy karyotype using the RT-PCR technique. Differential expression of these genes is quantified individually against a beta-Actin gene that has been employed as an internal control. The immunoblotting of cell lysate proteins with polyclonal antibodies exhibit SOD1 (16 kD), SOD2 (40 kD), GPx (23 and 92 kD), CAT (64 kD), and Actin (43 kD) as translational products. The results demonstrate that the enhancement in the level of mRNAs encoding SOD1 in DS male and female, as well as aged male and female are 51, 21, 31 and 50% respectively compared to the normal child (control). In SOD2, DS male and female display higher (176%) and lower (26%) levels of expression whereas aged male and female exhibit enhanced levels of expression (66 and 119%) respectively compared to the control. This study demonstrates that DS affects the female less than the male whereas in the aging process, the female is more prone to oxidative damage than the male. These results not only indicate that the level of GPx mRNA is constant except in DS male, which shows a downward regulation but that even CAT mRNA is upward regulated in aged as well as in DS males and females. These disproportionate changes in anti-oxidant genes, which are incapable of coping with over expressed genes, may contribute towards the aging process, dementia and Down syndrome.  相似文献   

20.
The effect of various doses (0, 10, 20, 40, or 80 mg/kg body weight) of naringin (a citrus flavonone) was studied on streptozotocin (STZ)-induced hyperglycaemic rats to evaluate the possible hypoglycaemic and antioxidant activity of naringin in diabetes. In comparison to the normoglycaemic group the treatment of rats with a single dose of STZ (65 mg/kg body weight) only revealed a significant increase (P < 0.05) in plasma hydrogen peroxide (H2O2) by 230%, increased the thiobarbituric acid reactive substances (TBARS) as index of the lipid peroxidation level by 69%, while total antioxidant activity was decreased by 36%, with a consistent significant decrease (P < 0.05) in the activity of erythrocytes antioxidative enzymes catalase (CAT), superoxide dismutase (SOD), glutathione peroxidase (GPx), and paraoxonase (PON). Exogenous administration of individual gradual doses of naringin to hyperglycaemic rats causes a dose-dependent decrease of the glucose level, an increase of the insulin concentration, a decrease of the H2O2 and TBARS levels, as well as the increase of the total antioxidant status with an increase of antioxidant enzyme activities (CAT, SOD, GPx, and PON). From this study, it may be concluded that all doses of naringin provided a significant amelioration of hypoglycaemic and antioxidant activity in STZ-induced diabetic rats, however, the greatest effect of naringin was observed at 80 mg/kg body weight.  相似文献   

设为首页 | 免责声明 | 关于勤云 | 加入收藏

Copyright©北京勤云科技发展有限公司  京ICP备09084417号