2D-DIGE-HPLC-nESIMS/MS对2,4-二硝基苯磺酸损伤HaCaT细胞差异蛋白质的鉴定

采用Cy2、Cy3和Cy5荧光染料标记蛋白,建立了人角质形成细胞HaCaT受2,4-二硝基苯磺酸(DNBS)刺激前后的双向胶内差异凝胶电泳(2D-DIGE)图谱,每组平行样本数为3。凝胶采用蛋白荧光染料Deep Purple进行后染色(Post-stain)。DeCyder定量分析软件在每块凝胶上平均检测到1 200个以上蛋白斑点,每块胶上都匹配得到的相同蛋白质斑点有846个。其中有7个斑点丰度变化在50%以上,统计学意义显著(P值小于0.05)。利用高效液相色谱-电喷雾串联质谱(HPLC-nESI MS/MS)成功鉴定5个表达上调的斑点分别为X染色体开放阅读框26(Cxorf26)、人辅分子伴侣23(PTGES3)、钙调蛋白(CALM3)、肌球蛋白轻链6(MYL6)和断裂点丛集区蛋白1(BANF1);2个表达下调蛋白斑点被鉴定为转录延伸因子B肽链2(TCEB2)和核糖体蛋白L23(RPL23)。除MYL6被报道与皮肤疾病相关外,其它蛋白与皮肤病变的关系有待研究。该研究得到的7个差异表达蛋白为DNBS类化学致癌物职业接触者皮肤病变研究提供了有价值的线索。     

维拉帕米作用下急性心梗大鼠比较蛋白质组学研究

以急性心梗大鼠为研究对象, 应用双向凝胶电泳法(2-DE)分析比较了维拉帕米作用下急性心梗大鼠心肌蛋白表达的差异, 从蛋白质水平探讨了维拉帕米心肌保护作用的发生机制. 结果表明, 与假手术组及模型组相比, 维拉帕米给药组心肌组织中有8个蛋白点表达显著上调, 7个蛋白点表达显著下调. 采用质谱(MALDI-TOF-MS)分析结合数据库检索, 共鉴定了其中的15种蛋白质, 可按功能分为如下4类: (1) 能量代谢及线粒体功能相关蛋白; (2) 氧化应激相关蛋白; (3) 细胞骨架蛋白; (4) 其它蛋白. 研究结果表明, 维拉帕米的心肌保护作用与恢复心肌损伤过程中的能量供应及对抗氧化应激等作用有关.     

中波紫外线与化学致癌物诱导下HaCaT细胞的蛋白表达应答

对角质形成细胞HaCaT分别进行中波紫外线(UVB)照射、2,4-二硝基苯磺酸(DNBS)刺激及UVB+DNBS(UD)共同刺激, 利用二维荧光差异胶内凝胶电泳(2D DIGE)、DeCyder定量分析软件和HPLC-nESI-MS/MS分析技术, 对HaCaT细胞产生的差异表达蛋白进行了鉴定. 有65个蛋白质点发生了明显表达差异(P<0.05), 与UVB或DNBS单独处理细胞相比, 有41个蛋白点表现为UVB和DNBS的正协同效应, 13个蛋白点表现为负协同效应, 5个蛋白点与UVB单独处理相近, 6个蛋白点与DNBS单独处理相近. HPLC-nESI-MS/MS从65个差异表达蛋白质点中共鉴定出60种单一(Unique)蛋白. 采用生物信息学方法对这些鉴定蛋白所涉及的分子功能、参与的生物学过程及信号通路进行了系统分析. 实验得到了与紫外辐射和化学诱导损伤的直接相关蛋白, 有助于研究不同环境条件下皮肤癌的形成及皮肤疾病的有效防护与治疗.     

串联飞行时间质谱鉴定急性脊髓损伤大鼠针刺后的差异蛋白质

针灸作为一种传统的康复治疗手段在治疗脊髓损伤方面已取得了较好疗效[1].本实验建立了急性脊髓损伤双向凝胶电泳图谱,初步观察了针刺治疗对急性脊髓损伤组织中蛋白质的变化与差异表达,用2种质谱技术鉴定了急性损伤相关差异蛋白质,对进一步从蛋白质组学角度研究脊髓损伤的病理机制和针刺治疗急性脊髓损伤的机理提供了重要参考数据.     

在镉盐胁迫下扇贝鳃组织应激蛋白的研究

采用透射电子显微镜观察了虾夷盘扇贝(Patinopecten yessoensis)鳃组织细胞的超微结构, 发现镉盐能胁迫鳃组织中的腮丝、细胞核和线粒体产生病变. 利用双向凝胶电泳(2D-PAGE)优化分离扇贝鳃组织的全蛋白, 获得约800个蛋白质斑点, 并筛选出37个由于镉盐胁迫而产生的差异蛋白质斑点. 选用基质辅助激光解吸离子化-飞行时间质谱(MALDI-TOF MS)技术和数据库检索鉴定差异蛋白, 结果发现7个与镉毒性密切相关的蛋白质, 即热休克蛋白70和β-淀粉酶等上调蛋白质及原肌球蛋白、肌动蛋白和钙活化核苷酸酶1等下调蛋白质. 此外, 还发现转录调节子Crp/Fnr家族为低表达蛋白质, 而ABC转运子为高表达蛋白质. 在这些差异蛋白中, 部分蛋白质适合作为连续监测流动海水中镉污染程度及评价其危害性的蛋白指示物.     

K-Means聚类分析法筛选柠檬香茅茎叶差异蛋白及鉴定

柠檬香茅含有大量的香茅精油,运用十分广泛,然而其茎、叶的精油含量却相差悬殊。 为探索柠檬香茅精油代谢相关的蛋白途径,本文对柠檬香茅旗叶、成熟叶及茎秆等材料进行精油含量、总蛋白含量测定及双向凝胶电泳(2-DE)表达谱分析,运用k-means聚类分析方法对2-DE电泳中差异蛋白斑点的丰度、等电点和相对分子质量进行聚类分析和讨论,结果表明,旗叶和茎秆上调表达的蛋白质斑点的聚类对于相对分子质量变化敏感,成熟叶上调表达蛋白质斑点对于丰度的变化较为敏感。 预测了精油代谢功能相关的蛋白质斑点15个,挖取预测蛋白质斑点通过基质辅助激光解吸电离飞行时间质谱(MALDI-TOF/TOF-MS)成功鉴定了9个蛋白质。 本研究为柠檬香茅精油的蛋白代谢途径提供新的基础信息及研究思路。     

蛋白质组学技术筛选与鉴定镉诱导下褐云玛瑙螺肝脏的差异蛋白质

比较丙酮/TCA沉淀法和直接裂解法,优化提取与分离褐云玛瑙螺(Achatina fulica, AF)肝脏全蛋白.采用丙酮/TCA沉淀法,可获得约600个蛋白质斑点.用0.5 mg/L CdCl2溶液浸泡后的去梗小白菜喂养AF,并作为镉盐诱导AF肝脏表达应激蛋白质的实验材料.采用蛋白质组学技术筛选出由镉盐诱导AF肝脏表达的14个差异蛋白质斑点,并用肽质量指纹图谱技术(peptide mass fingerprinting, PMF)和数据库比对法初步鉴定出7种差异蛋白质,其中部分为热激蛋白(heat shock protein)、甲基转移酶(Methyltransferase)、三磷酸腺苷结合盒子转运体(ABC transporter)、钼酸盐转运子亚基(molybdate transporter subunit)和磷酸甘油酸变位酶(phosphoglycerate mutase).差异蛋白质均参与镉盐代谢,并适合作为监测土壤或食物中镉污染程度及危害性的指示蛋白质.     

肺鳞癌和小细胞肺癌组织比较蛋白质组学研究

通过16例人肺鳞癌和小细胞肺癌组织中表达蛋白的二维电泳分离和质谱分析,经数据库检索鉴定了53个蛋白,其中24个蛋白与肺癌发病机制相关,4个蛋白在其它癌症中有报道.表达呈现差异的蛋白点有44个,其中34个在表达量上有差异,10个蛋白在鳞癌和小细胞癌间表现为有和无的关系.蛋白功能分析提示人肺鳞癌与小细胞癌的蛋白质组表达存在差异,分析这些差异蛋白有利于肺癌分型及其生物标志物研究.     

蛋白质组学技术筛选与鉴定鲤鱼嗅脑急性创伤后的应激蛋白质

优化建立鲤鱼(cyprinus carpio,CC)嗅觉端脑(嗅脑)全蛋白提取技术。联用低渗透裂解和液氮冻溶法破碎鲤鱼嗅脑组织(rhinencephalon tissue of cyprinus carpio,CCRT)、低速离心提取CCRT全蛋白,并采用双向凝胶电泳(2D-PAGE)技术进行有效分离。经分析与统计,每张CCRT的2D-PAGE图谱中的蛋白质斑点数目约为1200个。分别分离CCRT的脂溶性和水溶性全蛋白,并获得高分辨率的2D-PAGE图谱。选用差异蛋白质组学技术筛选经10%冰醋酸创伤后的CC,其端脑组织所表达出的6种应激蛋白质,并用肽质量指纹谱(peptide mass fingerprinting,PMF)和数据库检索技术给予鉴定。其中3种蛋白质为70S热休克蛋白、β微管蛋白和DNA链接酶IV,有望作为研究大脑急性创伤后的应激修复途径和机理的指示蛋白质。     

鼠肝癌淋巴道转移细胞模型的蛋白质组学研究

对2株来源于同一亲本细胞但淋巴道转移力显著不同的小鼠肝癌腹水型细胞株Hca-F(淋巴结转移率75%)和Hca-P(淋巴结转移率25%), 采用荧光差异双向凝胶电泳(2D DIGE)和DeCyder定量分析软件及HPLC-nESI-MS/MS技术, 定量分析和鉴定了小鼠肝癌细胞Hca-F和Hca-P的差异表达蛋白. 结果显示, 有116个蛋白质点表达水平存在明显差异(p<0.05), 在Hca-F中表达上调蛋白质点62个, 下调蛋白质点54个. 对所有116个蛋白质点进行了电喷雾串联质谱鉴定, 共鉴定出109种单一(Unique)蛋白. 其中部分蛋白已被报道与不同类型肿瘤的发生、浸润和转移相关, 多数蛋白质被首次报道与肝癌的淋巴道转移过程直接相关.     

Synthesis and structure of hydroxyisoxazolidines and derivatives of hydroxylamine and alkenals

The reactions of N-substituted hydroxylamines with alkenals serve as a method for the synthesis of the corresponding 2-substituted 3(5)-hydroxyisoxazolidines. The reaction pathway is determined by the nature of the substituent attached to the nitrogen atom. Ring-chain isomerism has been detected in these newly obtained compoundsTranslated from Khimiya Geterotsiklicheskikh Soedinenii, No. 9, pp. 1270–1276, September, 1987.     

Synthesis and characterization of triazenide and triazene complexes of ruthenium and osmium

Triazenide [M(eta2-1,3-ArNNNAr)P4]BPh4 [M = Ru, Os; Ar = Ph, p-tolyl; P = P(OMe)3, P(OEt)3, PPh(OEt)2] complexes were prepared by allowing triflate [M(kappa2-OTf)P4]OTf species to react first with 1,3-ArN=NN(H)Ar triazene and then with an excess of triethylamine. Alternatively, ruthenium triazenide [Ru(eta2-1,3-ArNNNAr)P4]BPh4 derivatives were obtained by reacting hydride [RuH(eta2-H2)P4]+ and RuH(kappa1-OTf)P4 compounds with 1,3-diaryltriazene. The complexes were characterized by spectroscopy and X-ray crystallography of the [Ru(eta2-1,3-PhNNNPh){P(OEt)3}4]BPh4 derivative. Hydride triazene [OsH(eta1-1,3-ArN=NN(H)Ar)P4]BPh4 [P = P(OEt)3, PPh(OEt)2; Ar = Ph, p-tolyl] and [RuH{eta1-1,3-p-tolyl-N=NN(H)-p-tolyl}{PPh(OEt)2}4]BPh4 derivatives were prepared by allowing kappa1-triflate MH(kappa1-OTf)P4 to react with 1,3-diaryltriazene. The [Os(kappa1-OTf){eta1-1,3-PhN=NN(H)Ph}{P(OEt)3}4]BPh4 intermediate was also obtained. Variable-temperature NMR studies were carried out using 15N-labeled triazene complexes prepared from the 1,3-Ph15N=N15N(H)Ph ligand. Osmium dihydrogen [OsH(eta2-H2)P4]BPh4 complexes [P = P(OEt)3, PPh(OEt)2] react with 1,3-ArN=NN(H)Ar triazene to give the hydride-diazene [OsH(ArN=NH)P4]BPh4 derivatives. The X-ray crystal structure determination of the [OsH(PhN=NH){PPh(OEt)2}4]BPh4 complex is reported. A reaction path to explain the formation of the diazene complexes is also reported.     

Preparation and characterization of diarylphosphazene and diarylphosphinohydrazide complexes of titanium, tungsten and ruthenium and phosphorylketimido complexes of rhenium

Reaction of the proligand Ph2PN(SiMe3)2 (L1) with WCl6 gives the oligomeric phosphazene complex [WCl4(NPPh2)]n, 1 and subsequent reaction with PMe2Ph or NBu4Cl gives [WCl4(NPPh2)(PMe2Ph)] (2) or [WCl5(NPPh2)][NBu4] (3), respectively. DF calculations on [WCl5(NPPh2)][NBu4] show a W=N double bond (1.756 A) and a P-N bond distance of 1.701 A, which combined with the geometry about the P atom suggests, there is no P-N multiple bonding. Reaction of L1 with [ReOX3(PPh3)2] in MeCN (X = Cl or Br) gives [ReX2(NC(CH3)P(O)Ph2)(MeCN)(PPh3)](X = Cl, 4, X = Br, 5) which contains the new phosphorylketimido ligand. It is bound to the rhenium centre with a virtually linear Re-N-C arrangement (Re-N-C angle = 176.6 degrees, when X = Cl) and there is multiple bonding between Re and N (Re-N = 1.809(7) A when X = Cl). The proligand Ph2PNHNMe2(L2H) reacts with [(C5H5)TiCl3] to give [(C5H5)TiCl2(Me2NNPPh2)] (6). An X-ray crystal structure of the complex shows the ligand (L2) is bound by both nitrogen atoms. Reaction of the proligands Ph2PNHNR2[R2 = Me2 (L2H), -(CH2CH2)2NCH3 (L3H), (CH2CH2)2CH2 (L4H)] with [{RuCl(mu-Cl)(eta6-p-MeC6H4iPr)}2] gave [RuCl2(eta6-p-MeC6H4iPr)L] {L = L2H (7), L3H (8), L4H (9)}. The X-ray crystal structures of 7-9 confirmed that the phosphinohydrazine ligand is neutral and bound via the phosphorus only. Reaction of complexes 7-9 with AgBF4 resulted in chloride ion abstraction and the formation of the cationic species [RuCl(6-p-MeC6H4iPr)(L)]+ BF4- {(L = L2H (10), L3H (11), L4H (12)}. Finally, reaction of complex 6 with [{RuCl(mu-Cl)(eta6-p-MeC6H4iPr)}2] gave the binuclear species [(eta6-p-MeC6H4iPr)Cl2Ru(mu2,eta3-Ph2PNNMe2)TiCl2(C5H5)], 13.     

Ti-V基储氢合金及其氢化物的物相结构与组分优化设计

Ti-V基储氢合金在室温、常压下即可表现出良好的储氢特性,且质量储氢容量明显高于传统AB₅型储氢合金,从而在氢气的精制和回收、运输和储存及热泵等方面有较早的应用。 此外,在混合气体分离、核反应堆中处理氢的同位素、镍氢电池及燃料电池负极材料等方面也得到了广泛的研究与关注。 基于目前Ti-V基储氢合金的研究现状,概述了该类合金的优势、限制性因素(包括成因)及改性手段。 此外,为了进一步理解Ti-V基合金储氢机理、构建合金组分与储氢特性之间的对应关系,本工作重点围绕Ti-V基储氢合金及其氢化物的结构、组分优化设计展开综述,并对其未来研究方向做出展望。     

Mass and NMR spectra of haplophyllidine and perforine and of their derivatives

Conclusions The mass and NMR spectra of haplophyllidine, perforine, and their derivatives have been studied. The influence of the open and cyclic forms of the molecular ion on the nature of the fragmentation has been discussed. The main routes of fragmentation of the compounds considered are due to the presence of substituents at C₈ and C₄.Khimiya Prirodnykh Soedinenii, Vol. 5, No. 4, pp. 273–279, 1969     

Investigation of adhesion and mobility of polyurethane and epoxy-rubber glues and adhesives

The values of activation parameters in uncured and cured epoxy resins, rubbers, and blends thereof are investigated. The dependences of activation energy and adhesion strength of epoxy-rubber compositions on rubber content are determined. The correlation of adhesion and activation energy values for polyurethane rubber and epoxy-rubber compositions is shown.     

Crystal and molecular structure of aroyl- and acetylhydrazones of acet- and benzaldehydes

Aroyl- and acetylhydrazones of acet- (I) and benzaldehydes (IV) and benzoylhydrazones of acet- (II) and benzaldehydes (III) were studied by x-ray structural and quantum-chemical methods in order to establish their structures. Compund (I) was the EEZ structure in the crystal. Calculations and spectral data showed that the EEE form occurs in nonpolar solvents and in the gas phase. According to crystallographic data molecules (I)–(IV) are the E-isomers (relative to the N-N bond) and the hydrazone fragments are planar. Intermolecular N-H...O H-bonds from in the crystals. The data obtained suggest that the majority of acylhydrazones are conformationally rigid on dissolution although exceptions do occur. Apparently the reasons for the difference of acetyl- and benzoylhydrazones in electrocarboxylation reactions are electronic and not steric factors.Translated from Izvestiya Akademii Nauk SSSR, Seriya Khimicheskaya, No. 1, pp. 75–81, January, 1991.     

Kinetics and mechanisms of chlorine dioxide and chlorite oxidations of cysteine and glutathione

Chlorine dioxide oxidation of cysteine (CSH) is investigated under pseudo-first-order conditions (with excess CSH) in buffered aqueous solutions, p[H+] 2.7-9.5 at 25.0 degrees C. The rates of chlorine dioxide decay are first order in both ClO2 and CSH concentrations and increase rapidly as the pH increases. The proposed mechanism is an electron transfer from CS- to ClO2 (1.03 x 10(8) M(-1) s(-1)) with a subsequent rapid reaction of the CS* radical and a second ClO2 to form a cysteinyl-ClO2 adduct (CSOClO). This highly reactive adduct decays via two pathways. In acidic solutions, it hydrolyzes to give CSO(2)H (sulfinic acid) and HOCl, which in turn rapidly react to form CSO3H (cysteic acid) and Cl-. As the pH increases, the (CSOClO) adduct reacts with CS- by a second pathway to form cystine (CSSC) and chlorite ion (ClO2-). The reaction stoichiometry changes from 6 ClO2:5 CSH at low pH to 2 ClO2:10 CSH at high pH. The ClO2 oxidation of glutathione anion (GS-) is also rapid with a second-order rate constant of 1.40 x 10(8) M(-1) s(-1). The reaction of ClO2 with CSSC is 7 orders of magnitude slower than the corresponding reaction with cysteinyl anion (CS-) at pH 6.7. Chlorite ion reacts with CSH; however, at p[H+] 6.7, the observed rate of this reaction is slower than the ClO2/CSH reaction by 6 orders of magnitude. Chlorite ion oxidizes CSH while being reduced to HOCl, which in turn reacts rapidly with CSH to form Cl-. The reaction products are CSSC and CSO3H with a pH-dependent distribution similar to the ClO2/CSH system.