Presence of natural anti-Galα1-4GalNAcβ1-3Gal (anti-NOR) antibodies in animal sera

Rare polyagglutinable NOR erythrocytes contain unusual globoside extention products terminating with a Galα1-4GalNAcβ1-3Gal- unit. This trisaccharide epitope is recognized by recently characterized antibodies naturally occurring in most human sera (Duk et al., Glycobiology, 15, 109, 2005). These antibodies represent two major types of fine specificity. All these antibodies are most strongly inhibited by Galα1-4GalNAcβ1-3Gal (NOR-tri), and weakly by Galα1-4Gal. However, the type 1 antibodies are strongly inhibited by Galα1-4Galβ1-3Gal-R and weakly by Galα1-4GalNAc, while the type 2 antibodies show the opposite reactivities with these two oligosaccharides. Similar antibodies have now been found in horse, rabbit and pig sera. The antibodies were purified from animal sera by affinity chromatography on Galα1-4GalNAcβ1-3Gal-human serum albumin(HSA)-Sepharose 4B conjugate. The specificity of the antibodies was determined by binding to ELISA plates coated with several α-galactosylated oligosaccharide-polyacrylamide (PAA) or -HSA conjugates and by inhibition with synthetic oligosaccharides. The purified antibodies bound specifically to conjugates containing NOR-tri. The inhibition of binding showed that the animal sera also contain two types of anti-NOR antibodies: type 2 was found in the horse serum, and a mixture of both types was present in rabbit and pig serum. These results indicate that anti-NOR, a new and distinct kind of anti-αGal antibody, are present in animal sera and show similar specificties and diversity as their counterparts found in human sera.     

Monoclonal antibodies that recognize the trisaccharide epitope Galα1-3Galβ1-4GlcNAc present on Ehrlich tumor cell membrane glycoproteins

Monoclonal antibodies were prepared against the trisaccharide Gal1-3Gal1-4GlcNAc, a sequence which occurs on the surface of Ehrlich ascites tumor cells as well as in thyroglobulin, laminin and a variety of other proteins. This was accomplished by immunizing BALB/c mice with the fraction of Ehrlich cell membrane glycoproteins obtained by affinity chromatography on aGriffonia simplicifolia I (GS I) column which selectively binds -d-galactosyl-terminated structures. Detection of Gal1-3Gal1-4GlcNAc-specific antibodies was accomplished by employing glycoproteins containing the trisaccharide sequence; fusion with spleen cells from an immunized mouse was accomplished in the presence of polyethylene glycol (PEG1500). An enzyme-linked immunosorbent assay (ELISA) system was used to identify two clones (2.10G and 6.8E), which recognized the desired trisaccharide conjugate. These clones also recognized a thyroglobulin fraction isolated by GS I affinity chromatography and murine laminin, both of which possess the Gal1-3Gal1-4GlcNAc sequence. Inhibition of antibody-trisaccharide reactivity, examined employing an ELISA assay, revealed that two trisaccharides, Gal1-3Gal1-4GlcNAc/Glc, were the best inhibitory haptens; Gal1-4GlcNAc (LacNAc), Gal1-3Gal and Gal1-4Glc (lactose) were poor inhibitors. Indirect immunofluorescence staining of unfixed Ehrlich cells using the monoclonal antibody at 4° C revealed fluorescence over the entire cell surface. Indirect immunogold labeling of semithin and ultrathin sections of aldehyde fixed and Lowicryl K4M-embedded Ehrlich cells resulted in specific labeling of the cell surface and internal structure. Immunoblot analysis revealed that removal of the -galactosyl residues of laminin by -galactosidase abolished reactivity with the monoclonal antibodies. The availability of this antibody, which belongs to the IgM family of immunoglobulins, now makes possible the detection of this sugar sequence on cells and tissue sections, as well as on glycoproteins in solution.     

α1-抗胰蛋白酶的研究进展

自1963年首次报道α1-抗胰蛋白酶(α1-AT)缺失与肺气肿的关系以来,α1-AT的结构和功能、作用原理等得到进一步的认识。伴随α1-AT产品的不断研制,对其临床应用及其临床研究更加广泛深入,对此进行综述。     

α1-抗胰蛋白酶的分离纯化

为建立从Cohn's组分IV沉淀中分离纯化α1-抗胰蛋白酶(α1-antitrypsin,α1-AT)的技术路线,以Cohn's组分IV为原料,利用还原剂1,4-二硫苏糖醇(DTT)、气相二氧化硅等处理,再经压滤、离子交换层析和疏水层析提取α1-AT.用SDS-PAGE及合成基质法检测α1-AT的纯度以及生物活性.结果可见,Cohn组分IV沉淀中α1-AT占蛋白总量的11%～13%,经该工艺提取的α1-抗胰蛋白酶纯度为97.6%,疏水层析后α1-AT的收获率为21%,比活为每毫克总蛋白中含 0.54 mg功能活性α1-AT.可见用此方法从Cohn组分IV沉淀中可制备高纯度的α1-AT.     

Phage antibodies: will new 'coliclonal' antibodies replace monoclonal antibodies?

Antibodies can now be rapidly isolated from large and diverse recombinant libraries by displaying functional antibody fragments on the surface of bacteriophage particles and directly selecting with antigen. This method has been used to isolate antibodies, including human antibodies, with and without immunization, and to improve the affinity and specificity of antigen binding.     

α1-抗胰蛋白酶的缺乏与治疗

α1-抗胰蛋白酶缺乏症(α1-antitrypsin deficiency,AATD)是遗传性疾病,可引起肝脏疾病,也与早发型肺气肿等肺部疾病有关。编码α1-抗胰蛋白酶(α1-antitrypsin,AAT)的基因具有复杂的单核苷酸多态性,并以常染色体共显性方式遗传。临床上,以混合人血浆为来源的静注AAT可提高AATD患者的血清AAT水平。除这种增补疗法外,新的治疗方法也在不断发展,包括基因治疗及干细胞治疗等。     

干扰素-α1b单用及联合胸腺肽α1治疗慢性乙型肝炎疗效观察

我院自1997年初开始使用胸腺肽α1(Tα1)联合干扰素-α1b(IFN-α1 b)治疗慢性乙型肝炎(CHB)患者,取得满意疗效,现报告如下.     

Comparison of the properties of concanavalin A and anti-α-d-glucose antibodies

Concanavalin A and anti-α-d-glucose antibodies form precipitin complexes with antigens havingα-d-glucose as terminal units. The sedimentation rates, molecular weights, gel electrophoretic mobilities, isoelectric points, and immunoglobulin type of Con A andα-Ab have been determined. The interactions of the compounds with antigens in the presence of potential inhibitors have been compared. The data show that the interaction of Con A with glucose units occurs with hydrogen bonding at hydroxyl groups at C1, 3,4, and 6 and van der Waals bonding at the pyranose ring oxygen. In theα-Ab complex with glucose units, in addition to the above bond types, a hydrogen bond at the hydroxyl at C2 occurs and this bond is essential for interaction.     

α-1-酸性糖蛋白1在肺癌中的表达及其生物学功能探究

α-1-酸性糖蛋白(alpha-1-acid glycoprotein 1, AGP1)属于orosomucoid家族成员,主要由肝细胞和中性粒细胞分泌,是相对分子质量为23.51 kD的酸性糖蛋白。前期研究发现, AGP1蛋白在早期非小细胞肺癌(non-small cell lung cancer, NSCLC)患者血清中含量下降,有望作为NSCLC早诊标志物,但其在肺癌组织中的表达、具体生物学作用及临床意义尚不清楚。本研究运用GEPIA数据库分析肺癌组织中AGP1 m RNA的表达水平,采用蛋白质印迹法检测AGP1蛋白在肺癌组织和血清中的含量,利用cBioPortal数据库分析AGP1在肺癌组织中的DNA改变频率,利用STRING数据库构建与AGP1蛋白相互作用的蛋白质网络,利用HPA和Kaplan-Meier数据库分析AGP1的表达水平与肺癌患者生存的关系;同时,构建过表达AGP1基因的A549和H157肺癌细胞系,通过CCK-8实验检测过表达AGP1对A549和H157细胞增殖的影响。结果显示:AGP1的m RNA水平在486例肺鳞癌组织中显著下降,在早期肺癌组织和血清中AGP...     

Synthesis of p-trifluoroacetamidophenyl O-α-D-galactopyranosyl-(1→2)-O-α-D-mannopyranosyl-(1→4)-α-L-rhamnopyranoside

The role of exposed tyrosine side-chains in enzyme-catalysed reactions has been studied for porcine-pancreatic alpha-amylase, sweet-potato beta-amylase, and Aspergillus niger glucamylase using N-acetylimidazole as the specific protein reagent. The changes in activity binding affinity (Δk_?1/k₊₁), and kinetic parameters (K_m,k₂) due to acetylation of the phenolic hydroxyl groups have been determined. Acetylation of each enzyme occurred by an “apparent” first-order reaction with a rate constant of 0.72–1.4 x 10^?1min^?1. Acetylation increased the apparent K_m (soluble starch as the substrate) for each enzyme (appreciably for alpha-amylase and glucamylase), whereas k² remained unchanged. Similarly, for each enzyme, the binding affinity for immobilised cyclohexa-amylose decreased appreciably, whereas the catalytic activity was reduced only to a small degree (and remained unchanged for beta-amylase). It is concluded that the tyrosine groups located in the active centre of each enzyme have a substrate-binding function.     

微生物学方法制备16α-甲基-11α，17α，21-三羟基孕甾-1，4-二烯3，20-二酮

本文报道了简单节杆菌A69-2和球孢白僵菌AS69同时存在下对16α-甲基-17α-羟基孕甾-4．烯-3,20-二酮-21-醋酸酯(16MRSA)的协周转化作用。 这种协同转化怍用既能解除16α-甲基-11α,17a,21-三羟基孕甾-4-烯-3,20-二酮(16MllaHC)对球孢白僵菌AS69的11α羟化酶的抑制作用,又可降低高浓度的16M11aHC对节杆菌A69—2脱氢酶活性的影响,同时还能抑制节杆菌脱氢过程的副反应。在底物浓度为0.15％(W／V)时,l6α-甲基-11α,17a,21-三羟基孕甾-1,4-二烯-3,20-二酮(16MDHC)的收率约50%,故是制备1 6MDHC的一种理想的微生物学方法。     

α-环糊精葡萄糖基转移酶催化合成α-熊果苷

以对苯二酚和麦芽糊精为底物,通过α-环糊精葡萄糖基转移酶和淀粉葡萄糖苷酶的两步酶法反应体系催化合成α-熊果苷。优化后的催化条件:以葡萄糖当量(DE)值为8%~10%的麦芽糊精作为供体底物,麦芽糊精60g/L,对苯二酚150 mmol/L,缓冲液p H 6.0,在40℃下反应24 h。在此反应条件下,α-熊果苷的产量为3.17 g/L,对苯二酚转化率为7.77%。通过萃取法对α-熊果苷进行了初步分离,再经高效液相色谱-电喷雾串联质谱技术进行了结构测定,确定产物为α-熊果苷。     

白细胞介素-1α的成熟与分泌机制

白细胞介素-1α(Interleukin-1α,IL-1α)是IL-1家族中重要的一员。IL-1α是广泛存在于机体中的多功能信号分子,在胞内作为重要的转录因子,调控细胞生长分化;同时可分泌至胞外参与机体炎症应答,在癌症等多种疾病的发生发展中发挥着重要的作用。IL-1α前体和成熟形式都具有生物活性,但成熟形式的IL-1α生物活性大大增强。IL-1α是许多疾病预防治疗的靶标,其成熟与分泌的机制也备受关注。近年的研究表明,病原体感染宿主诱导IL-1α的成熟与分泌受细胞内钙离子浓度及钙蛋白酶活性的调控,部分情况下还有炎症小体信号途径及其他信号途径等。本文将结合本课题组的研究成果对IL-1α的成熟与分泌机制及相关研究进行综合评述。     

人α-1干扰素cDNA全序列分析

1982年,我们用新城疫病毒F株诱生的人脐血白细胞mRNA逆转录成cDNA,建立了人α型干扰素基因无性繁殖系,经限制性内切酶谱分析,以及采用化学法就HaeⅢBglⅡ片     

AHR和HIF1-α代谢调控1型调节性T细胞分化

<正>目前,对于淋巴细胞代谢通路,及代谢本身和代谢产物对免疫反应的调节作用,仍不甚清楚。本文作者发现,缺氧诱导因子1-α(hypoxia inducible factor-1α,HIF1-α)和芳基化合物受体(aryl hydrocarbon receptor,AHR)通过代谢编码调控1型调节性T细胞(type 1 regulatory T cell,Tr1)分化。HIF1-α调节Tr1早期代谢重编码。随后,AHR促进HIF1-α降解,接管对T细胞代谢     

微生物合成11α-羟基-16α,17α-环氧孕酮的动力学

对耐温黑根霉菌丝体催化16α,17-α-氧孕酮生成11α-羟基-16α,17α-环氧孕酮的反应体系进行了研究,考察了不同反应时间、不同菌体量和底物质量浓度对11α-羟化反应的影响,建立了相应的动力学模型,其方程为r=0.031Sr/1.05＋Sr。结果表明：提高菌体质量浓度有利于提高反应速率;底物浓度与反应初速率的关系与米氏方程相似。     

Proteolytic inactivation of 1, 4-α-d-glucan 6-α-d-glucosyltransferase purified from Aspergillus niger R-27

An extracellular 1,4-α-d-glucan 6-α-d-glucosyltransferase [d-glucosyltransferase, 1,4-α-d-glucan:1,4-α-d-glucan(d-gluco 6-α-d-glucosyltransferase, EC 2.4.1.24] from Aspergillus niger R-27 has been purified and the kinetics of its proteolytic inactivation with subtilisin studied. The purified enzyme was shown to be homogeneous using disc polyacrylamide gel electrophoresis. It contained 16.0% mannose, 0.19% glucose and 2.95% 2-acetamido-2-deoxy-d-glucose. The characteristic feature of the proteolytic degradation of glucosyltransferase is rapid hydrolysis of ～12 peptide bonds per mol and the formation of an active intermediate product which is more resistant to further proteolysis, but is easily heat-inactivated. The isolation and some properties of glucosyltransferase are also described.