细叶百合Catalase基因的克隆及表达分析

为探究细叶百合(Lilium pumilum)Catalase(CAT)基因与耐盐碱胁迫的关系,从细叶百合鳞茎中成功克隆出LpCat基因。该基因的ORF区长度为1479 bp,编码492个氨基酸,对其进行序列比对和系统进化树分析,发现细叶百合Catalase蛋白与通江百合(L.sargentiae)、菠萝(Ananas comosus)、油棕(Elaeis guineensis)等植物的Catalase蛋白亲缘关系较近。构建了pQE-LpCat原核表达载体,以1 mol·L^(-1)IPTG为诱导剂进行LpCat蛋白的体外诱导表达与纯化。在添加50 mmol·L^(-1)NaHCO_(3)胁迫下,携带PQE-LpCat蛋白的菌液生长浓度高于对照菌株。在1 mol·L^(-1)NaHCO_(3)、2.5 mol·L^(-1)H_(2)O_(2)胁迫下过表达LpCAT基因烟草(Nicotiana plumbaginifolia)植株与野生型相比,枯萎程度相对较低。通过净光合速率、气孔导度、胞间CO_(2)和蒸腾速率,H_(2)O_(2)含量,丙二醛(MDA)等生理指标检测说明过表达LpCat基因烟草植株比野生型烟草植株更耐盐碱。

幽门螺杆菌Catalase基因的克隆、表达及其抗原性鉴定

目的：构建含人幽门螺杆菌（Hpylori，Hp）过氧化氢酶（catalase，KatA）编码基因的重组质粒，测定、分析其核酸序列，并在E.coli中表达，研究其抗原性。方法：应用PCR技术从HpDNA染色体中扩增KatA编码基因片段，将其T-A克隆和测序，并与GenBank公布的其他Hp菌株的基因序列比较，再将目的基因插入至融合表达载体pGEX-4T-1中进行表达，用GST亲和层析对其进行纯化。纯化产物用于对29株小鼠抗Hp-全菌单克隆抗体（mAb）的鉴定及与Hp啊感染患者血清进行Western blot。结果：KatA基因全长为1 515 bp，并在GenBank上登录（No．DQ333889），与GenBank公布的其他Hp菌株的核酸的同源性为96％～97％，表达的KatA融合蛋白的相对分子质量（Mr）为85 000，29株小鼠抗Hp全菌mAb中有4株mAb是针对KatA的，表达产物可被Hp感染患者的血清特异性识别。结论：重组KatA具有较好的抗原性，为坳检测试剂和疫苗的研究奠定了基础。

幽门杆菌Catalase/GST融合蛋白的表达、标签切除及鉴定

旨在利用GST融合基因表达系统表达幽门螺杆菌Catalase融合蛋白,并利用凝血酶切除GST标签。将重组质粒Catalase/pGEX-4T-1转化大肠杆菌BL21(DE3)感受态中,用IPTG进行诱导表达,菌体经反复冻融、溶菌酶裂解及超声破菌后,Catalase/GST融合蛋白以部分可溶性的形式表达在上清中。采用谷胱甘肽琼脂糖树脂Glutathione Sepharose 4B对其进行纯化,得到Catalase/GST融合蛋白,再用凝血酶进行GST标签的切除,所得产物进行Western blotting鉴定。高效表达出Catalase/GST融合蛋白的相对分子质量约85 kD,凝血酶成功地切除了GST标签,Western blotting证实Catalase蛋白能被鼠抗Catalase单克隆抗体识别。

Response of soil catalase activity to chromium contamination

The impact of chromium（Ⅲ） and （Ⅵ） forms on soil catalase activity was presented. The Orthic Podzol, Haplic Phaeozem and Mollic Gleysol from different depths were used in the experiment. The soil samples were amended with solution of Cr（Ⅲ） using CrCl3, and with Cr（Ⅵ） using K2Cr2O7 in the concentration range from 0 to 20 mg/kg, whereas the samples without the addition of chromium served as control. Catalase activity was assayed by one of the commonly used spectrophotometric methods. As it was demonstrated in the experiment, both Cr（Ⅲ） and Cr（Ⅵ） have an ability to reduce soil catalase activity. A chromium dosage of 20 mg/kg caused the inhibition of catalase activity and the corresponding contamination levels ranged from 75% to 92% for Cr（Ⅲ） and 68% to 76% for Cr（Ⅵ）, with relation to the control. Catalase activity reached maximum in the soil material from surface layers （0-25 cm）, typically characterized by the highest content of organic matter creating favorable conditions for microorganisms.

Modulatory effect of pineapple peel extract on lipid peroxidation,catalase activity and hepatic biomarker levels in blood plasma of alcoholinduced oxidative stressed rats

Objective:To investigate the ability of the methanolic extract of pineapple peel to modulate alcohol-induced lipid peroxidation,changes in catalase activities and hepatic biochemical marker levels in blood plasma.Methods:Oxidative stress was induced by oral administration of ethanol(20%w/v) at a dosage of 5 niL/kg bw in rats.After 28 days of treatment,the rats were fasted overnight and sacrificed by cervical dislocation.Blood was collected with a 2 mL syringe by cardiac puncture and was centrifuged at 3000 rpm for 10 min.The plasma was analyzed to evaluate malondialdehyde(MDA),catalase activity,aspartate aminotransferase(AST),alkaline phosphatase(ALP) and alanine aminotransferase(ALT) concentrations.Results:Administration of alcohol caused a drastic increase(87.74%) in MDA level compared with the control.Pineapple peel extract significantly reduced the MDA level by 60.16%at 2.S mL/kg bw.Rats fed alcohol only had the highest catalase activity,treatment with pineapple peel extract at 2.5 mL/kg bw however, reduced the activity.Increased AST,ALP and ALT activities were observed in rats fed alcohol only respectively,treatment with pineapple peel extract drastically reduced their activities. Conclusions:The positive modulation of lipid peroxidation,catalase activities as well as hepatic biomarker levels of blood plasma by the methanolic extract of pineapple peels under alcoholinduced oxidative stress is an indication of its protective ability in the management of alcoholinduced toxicity.

Cloning of catalase and expression patterns of catalase and selenium-dependent glutathione peroxidase from Exopalaemon carinicauda in response to low salinity stress

Catalase （CAT） and selenium-dependent glutathione peroxidase （Se-GPx） play a vital role in protecting organisms against various oxidative stresses by eliminating H202, The objective of this paper is to evaluate the roles of these antioxidant molecules in the ridgetail white prawn Exopalaemon carinicauda in response to low salinity stress. A complementary DNA （cDNA） containing the complete coding sequence of CAT was cloned from the hepatopancreas using reverse-transcription polymerase chain reaction （RT-PCR） and rapid amplification of cDNA ends. The full-length cDNA of CAT （2 649 bp） contains a 5＇-untranslated region （UTR） of 78 bp, a 3＇- UTR of 1 017 bp, with a poly （A） tail, and an open reading frame of 1 554 bp encoding a 517-amino-acid polypeptide with predicted molecular mass of 58.46 kDa and estimated isoelectric point of 6.64. This CAT sequence contained the proximal active site signature （60FDRERIPERWHAKGAG76）, proximal heme-ligand signature sequence （350RLFSYPDTH358） and three catalytic amino acid residues （His71, Asn144 and Tyr354）. Sequence comparison showed that the CAT deduced amino acid sequence of E. carinicauda shared 68%-92% of identities with those of other species. Quantitative real-time PCR analysis revealed that CAT mRNA was widely expressed in the hepatopancreas （highest）, hemocyte, eyestalk, heart, gill, muscle, ovary and stomach. Under low salinity stress, CAT and GPx mRNA expression levels both in the gill and hepatopancreas increased significantly at the first 48 h and 6 h respectively, indicating a tissue- and time-dependent antioxidant response in E. carinicauda. All these results indicate that E. carinicauda CAT is a member of the CAT family and might be involved in the acute response against low salinity stress.

Inflammation/bioenergetics-associated neurodegenerative pathologies and concomitant diseases:a role of mitochondria targeted catalase and xanthophylls

Various inflammatory stimuli are able to modify or even"re-program"the mitochondrial metabolism that results in generation of reactive oxygen species.In noncommunicable chronic diseases such as atherosclerosis and other cardiovascular pathologies,type 2 diabetes and metabolic syndrome,these modifications become systemic and are characterized by chronic inflammation and,in particular,"neuroinflammation"in the central nervous system.The processes associated with chronic inflammation are frequently grouped into"vicious circles"which are able to stimulate each other constantly amplifying the pathological events.These circles are evidently observed in Alzheimer's disease,atherosclerosis,type 2 diabetes,metabolic syndrome and,possibly,other associated pathologies.Furthermore,chronic inflammation in peripheral tissues is frequently concomitant to Alzheimer's disease.This is supposedly associated with some common genetic polymorphisms,for example,Apolipoprotein-Eε4 allele carriers with Alzheimer's disease can also develop atherosclerosis.Notably,in the transgenic mice expressing the recombinant mitochondria targeted catalase,that removes hydrogen peroxide from mitochondria,demonstrates the significant pathology amelioration and health improvements.In addition,the beneficial effects of some natural products from the xanthophyll family,astaxanthin and fucoxanthin,which are able to target the reactive oxygen species at cellular or mitochondrial membranes,have been demonstrated in both animal and human studies.We propose that the normalization of mitochondrial functions could play a key role in the treatment of neurodegenerative disorders and other noncommunicable diseases associated with chronic inflammation in ageing.Furthermore,some prospective drugs based on mitochondria targeted catalase or xanthophylls could be used as an effective treatment of these pathologies,especially at early stages of their development.

Association of Catalase Genotype with Oxidative Stress in the Predication of Colorectal Cancer:Modification by Epidemiological Factors

Objective This paper aims to assess the interaction between common variations in catalase（CAT） polymorphic gene and environmental factors for antioxidant defense enzyme in modulating individual susceptibility to colorectal cancer（CRC）.Methods A case-control study with 880 colorectal cancer cases and 848 controls was conducted to investigate whether variations in the catalase（CAT） gene,one of the genes involved in scavenging oxidative stress,influenced susceptibility to CRC.Results The interaction between life style and genotypes as well as with their effects on colorectal cancer was deduced from the present study.Significant difference（P=0.01） was identified in the distribution of CAT genotype between the colorectal cancer cases and the controls.The CRC cases had significantly lower mean activity than the controls（P〈0.01）.Correlation analyses revealed statistically significant correlations between CAT activity and CAT genotype（P〈0.01）.Conclusion The risk of CRC was associated with smoking,low vegetable consumption,high pork and poultry consumptions,and low or high BMI.This is the first study reporting an association of polymorphism CAT-21A〉T with colorectal cancer.Low CAT activity was associated with an increased risk of CRC;however,no evidence was found to support an association between CAT-21A〉T polymorphism and CRC risk.

HIGHLY SENSITIVE CATALASE ELECTRODE BASED ON POLYPYRROLE FILMS WITH MICROCONTAINERS

Highly sensitive catalase electrodes for sensing hydrogen peroxide have been fabricated based on polypyrrole films with microcontainers. The microcontainers have a cup-like morphology and are arranged in a density of 4000 units cm^-2. Catalase was immobilized into the polypyrrole films with microcontainers （Ppy-mc）, which were coated on a Pt substrate electrode. The catalase/Ppy-mc/Pt electrode showed linear response to hydrogen peroxide in the range of 0-18 mmol/L at a potential of-0.3 V （versus SCE）. Its sensitivity was measured to be approximately 3.64 μA （mmol/L） ^-1 cm^- 2, which is about two times that of the electrode fabricated from a flat Ppy film （catalase/Ppy-flat/Pt electrode）. The electrode is highly selective for hydrogen peroxide and its sensitivity is interfered by potential interferents such as ascorbic acid, urea and fructose. Furthermore, such catalase electrodes showed long-term storage stability of 15 days under dry conditions at 4℃.

Mailuoning suppresses H_2O_2-induced cortical neuronal injury and correlates with increased catalase activity and inhibited oxidative stress function

BACKGROUND：Mailuoning, a Chinese herb, has been widely used in China to treat acute ischemic stroke, and the major component exhibits anti-oxidative effects. However, the precise anti-oxidation pathway remains uncertain.OBJECTIVE：To validate the protective effects of Mailuoning on H202-induced primary cortical neuron injury in embryonic mice.DESIGN, TIME AND SETTING：Comparative observation and in v#ro experiments were performed at the Jiangsu Key Laboratory for Molecular Medicine from January 2008 to September 2009.MATERIALS：Mailuoning （Nanjing Jinling Medical Company, China）, reactive oxygen species （ROS） kit （Beyotime Biotechnology, China）, superoxide dismutase （SOD）, Cu/Zn SOD kit, malondialdehyde （MDA） kits （Nanjing Jiancheng, China）, mitochondrial membrane potential （GMS10013.1, GENMED, USA） and catalase activity assay kit （Beyotime Biotechnology, China） were utilized for the present study.METHODS：Mouse embryonic cortical neurons were isolated and cultured with culture medium containing H2O2 （80 μmol/L） and/or Mailuoning （1.25 μg/mL） for 24 hours.MAIN OUTCOME MEASURES：Neuronal viability and death were detected by methyl thiazolyl tetrazdium and flow cytometry; ROS production was determined by flow cytometry; mitochondriai membrane potential was detected using fluorescent staining; SOD activity was detected using a modified nitroblue tetrazolium method; Cu/Zn SOD and catalase activity was detected by spectrophotometry; and MDA was determined using the lipid peroxidation method.RESULTS：H2O2 increased ROS production and MDA concentration （P 〈 0.05）, and decreased mitochondrial membrane potential, SOD, Cu/Zn SOD and catalase activity （P 〈 0.05）; the number of surviving neurons （P 〈 0.05） was also reduced. Mai/uoning reversed these changes.CONCLUSION：Mailuoning protects H2O2-induced injury in cortical cells by inhibiting ROS and MDA, increasing depolarization of mitochondrial membrane, and enhancing SOD and catalase activity.

Glutathione peroxidase, superoxide dismutase and catalase activities in children with chronic hepatitis

The advantages of measuring hepatic oxidative status in liver biopsy are that it helps in diagnosis of hepatic dysfunction, reflects the degree of deterioration in the liver tissues, and helps to determine the severity of hepatic injury. We aimed to study the oxidative stress state in children with chronic hepatitis by using indirect approach in which antioxidant enzymes such as glutathione peroxidase (GPX), superoxide dismutase (SOD) and catalase (CAT) are determined in the liver tissue. The present study included 21 children and adolescents (12 males, 9 females) suffering from chronic hepatitis. Patients were selected from the Hepatology Clinic, New Children’s Hospital, Cairo University from November 2006 till 2009 and compared with a group of 7 children who happened to have incidental normal liver biopsy. Children with chronic hepatitis had mean age 8.12 ± 1.15 years. It was further subdivided into 2 subgroups: chronic viral heaptitis (n = 13) and cryptogenic hepatitis (n = 8). GPX, SOD and CAT levels were measured in fresh liver tissue (cell free homogenates) using ELISA. In chronic hepatitis group;there was a significant increase in the hepatic GPX activity (38.59 ± 35.82 nmol/min/ml) as compared to the control group (10.62 ± 6.68 nmol/min/ml). Also a significant correlation was observed between SOD and both ALT (r = 0.87, p < 0.05) and AST (r = 0.74, p < 0.05). GPX correlated with ALT (r = 0.80, p < 0.05) level in the chronic viral hepatitis subgroup. Our findings suggest that oxidative stress could play a role in the pathogenesis of chronic hepatitis. These preliminary results are encouraging to conduct more extensive clinical studies combining antioxidant therapy with various treatments.

Investigating Catalase and Carbonic Anhydrase Enzyme Activities and Levels of Certain Trace Elements and Heavy Metals in Patients with Primary and Metastatic Hepatic Carcinoma

Hepatocellular carcinoma (HCC) is among most common terminal cancer types in the world. Primary etiological factors include cirrhosis, hepatitis, aflatoxin and alcohol. The current study was conducted to determine cytosolic erythrocyte carbonic anhydrase and catalase enzyme activities and levels of some trace elements. For this purpose, 40 patients with primary and metastatic hepatic cancer and 29 healthy volunteers enrolled to the study. Catalase and carbonic anhydrase enzyme activities and serum trace element levels were measured in patient and control groups. In the current study, serum copper, magnesium, manganese and zinc levels were lower in the primary and metastatic hepatic cancer group in comparison with the control group (P < 0.05). In contrast, serum iron, cobalt, cadmium and lead levels were higher in the patient relative to the control group (P < 0.05). In comparison with the control group, the catalase level was lower in primary and metastatic cancer group, while the carbonic anhydrase level was higher in the cancer group (P < 0.05). Changes in levels of trace elements and anti-oxidant enzymes may be the factors which influence the development and progression of liver cancer. The carbonic anhydrase enzyme can be a useful indicator in the diagnosis of cancer. However, this issue warrants further investigation.

Effect of Petroleum Products on Soil Catalase and Dehydrogenase Activities

The effect of refined petroleum products on the activities of selected enzymes (catalase and dehydrogenase) was studied. There was a significant decrease (p kerosene > diesel > engine oil. However, a significant increase (p diesel > petrol > engine oil. On the whole the results reveal that refined petroleum products alter soil biochemistry.

Osmolyte modulated enhanced rice leaf catalase activity under salt-stress

Change in catalase activity was examined in leaves of rice plant exposed to salinity. Depending on the method of preparation of crude protein extract from leaf and the constituents of the assay medium, a significant difference in enzyme activity was recorded. Inclusion of sorbitol or mannitol or sucrose in the extraction and enzyme assay medium enhanced the enzyme activity in salt-stressed samples by nearly 1.5-1.8 fold, compared to the activity found in un- stressed plants, which otherwise showed a 50% declined activity in leaf extract prepared in buffer solution and assayed in a medium depleted of these sugars. In view of the accumulation of osmolytes under saline condition, these observations suggest that the catalase activity is modulated by the osmolytes and maintains a high rate of hydrogen peroxide scavenging property in vivo and serves as the major antioxidant enzyme to scavenge the salt-induced formation of peroxide. Therefore, the salt-stress induced appearance of low activity of the enzyme under normal buffer extraction and assay conditions, as reported in literature may represent an apparent than for its real in vivo activity.

精神分裂症患者与过氧化氢酶（Catalase）基因多态性

目的：探讨过氧化氢酶（Catalase）基因多态性与精神分裂症的相关性。方法：采用Amp-RFLP（amplication-restriction freg-ment lengh polymorphis）方法对精神分裂症患者和对照组的过氧化氢酶基因相关性进行了研究。结果：167例精神分裂症患者与155例对照组之间过氧化氢酶基因型频率和等位基因频率无统计学意义,76例女性精神分裂症患者与60例对照组之间过氧化氢酶基因型频率（x^2=11.096,df=2,P=0.004）有显著性差异。结论：过氧化氢酶多态性与精神分裂症的遗传学并不存在关联性,但是研究表明过氧化氢酶基因型频率与女性精神分裂症患者有显著性差异。

Molecular cloning, characterization and expression analysis of a catalase gene in Paphia textile

Catalase is an important antioxidant protein that can protect organisms against various forms of oxidative damage by eliminating hydrogen peroxide. In this study, the catalase c DNA of Paphia textile（Pt CAT） was cloned using RTPCR and rapid amplification of c DNA ends（RACE）. Pt CAT is 1 921 bp long and consists of a 5′-UTR of 50 bp, a 3′-UTR of 349 bp, and an ORF of 1 542 bp that encodes 513 amino acids with a molecular weight of 58.4 k D and an estimated isoelectric point of 8.2. Sequence alignment indicated that Pt CAT contained a highly conserved catalytic signature motif（^（61）FNRERIPERVVHAKGAG^（77））, a proximal heme-ligand signature sequence（^（352）RLFSYSDP^（359））, and three catalytic amino acid residues（H^（72）, N^（145）, and Y^（356））. Pt CAT also contains two putative N-glycosylation sites（^（34）NKT^（36） and ^（437）NFT^（439）） and a peroxisome-targeting signal（^（511）AQL^（513））. Furthermore, Pt CAT shares 53%–88% identity and 29%–89% similarity with other catalase amino acid sequences. Pt CAT m RNA was present in all tested organs, including the heart, digestive gland, adductor muscle, gonad, gill, and mantle, but its expression was highest in the digestive gland. High-temperature-induced stress produced two expression patterns of Pt CAT m RNA： first, an initial up-regulation followed by a down-regulation in the heart, digestive gland, and gonad and, second, consistent down-regulation in all other organs. These results demonstrate that Pt CAT is a typical member of the catalase family and might be involved in the responses to harmful environmental factors.

Molecular cloning, characterization, and antioxidant function of catalase in Lymantria dispar asiatic (Lepidoptera: Lymantriidae) under avermectin stress

The critical antioxidant catalase(CAT)breaks down hydrogen peroxide induced by environmental stresses.Here we cloned full length catalase cDNA from Lymantria dispar asiatic(LdCAT).Bioinformatic analyses showed that open reading frames of LdCAT contains 1524 bp,encoding 507 amino acids with molecular weight of 126.99 kDa,theoretical pI of 5.00,aliphatic index of 29.92,grand average of hydropathicity of 0.764,and instability index(II)of 46.56.Protein BLAST and multiple sequence alignment indicated that LdCAT had high identity with CAT from other insects,especially lepidopterans.In a phylogenetic analysis,LdCAT was most similar to CAT from Spodoptera litura and S.exigua.Quantitative realtime polymerase chain reaction showed that LdCAT transcripts in all instar larvae and the five tissues tested,verifying the ubiquity of LdCAT in L.disapr.Moreover,LdCAT of third instar larvae was significantly upregulated after they fed on avermectin at sublethal and LC10 doses.The highest relative transcript levels were found 2 h after an avermectin spray at LC90,and in the cuticula,rather than heads,fat bodies,malpighian tubes,and midguts after a spray avermectin at a sublethal concentration.The expression level of LdCAT under pesticide stresses here suggested that CAT is an important antioxidant enzyme of L.disapr defensing against pesticide stress and may be a good target for controlling this pest.

Effect of Catalase on Biocatalytic Synthesis of Pyruvate by Enzymes from Pseudomonas sp.

Pyruvate was produced from DL-lactate by a kind of green-chemical biocatalyst cell-free extract from bacterial strain Pseudomonas sp. SM-6. Catalase in cell-free extract, which could stabilize the pyruvate formed by lactate oxidase, played an important role in pyruvate preparation. The effect of catalase in conversion process was evaluated.

Direct Electrochemistry of Catalase on Single Wall Carbon Nanotubes Modified Glassy Carbon Electrode

Direct electrochemistry of catalase (Ct) has been studied on single wall carbon nanotubes (SWNTs) modified glassy carbon (GC) electrode. A pair of well-defined nearly reversible redox peaks is given at –0.48 V (vs. SCE) in 0.1 mol/L phosphate solution (pH 7.0). The peak current in cyclic voltammogram is proportional to the scan rate. The peak potential of catalase is shifted to more negative value when the pH increases. Catalase can adsorb on the SWNTs modified electrode.

Effect of N+ Beam Exposure on Superoxide Dismutase and Catalase Activities and Induction of Mn-SOD in Deinococcus Radiodurans

Though bacteria of the radiation-resistant Deinococcus radiodurans have a high resistance to the lethal and mutagenic effects of many DNA-damaging agents, the mechanisms involved in the response of these bacteria to oxidative stress are poorly understood. In this report, the superoxide dismutase (SOD) and catalase (CAT) activities produced by these bacteria were measured, and the change of SOD and CAT activities by 20 keV N+ beam exposure was examined. Their activities were increased by N+ beam exposure from 8x 1014 ions/cm2 to 6x1015 ions/cm2.The treatment of H2O2 and [ CHC13 +CH3 CH2OH ] and the measurement of absorption spectrum showed that the increase in SOD activity was resulted from inducible activities of MnSOD in D. radiodurans AS1.633 by N+ beam exposure . These results suggested that this bacteria possess inducible defense mechanisms against the deleterious effects of oxidization.