烟草与蓝猪耳胚胎发生中细胞表面三种凝集素受体的动态分布

以异硫氰酸甲酯(FITC)标记的三种凝集素(伴刀豆凝集素, 麦芽凝集素和大豆凝集素)为荧光探针,对烟草及蓝猪耳各发育时期胚细胞表面的凝集素受体进行了定位.结果显示胚柄基部荧光信号最强,沿胚柄单列细胞向胚体方向渐次减弱.以后随着胚柄功能的逐渐丧失而改变.同时,三种凝集素受体集中分布于胚柄细胞间的分裂面;凝集素受体在原胚中分布的另一个特点是聚集于新形成的细胞壁上.随着胚胎发育至分化阶段,凝集素受体则主要分布在胚体细胞的外切向壁上;三种凝集素受体的动态分布显示了凝集素受体的分布与细胞分裂之间的密切关系及其调控胚胎发育的作用.     

麦胚凝集素作用下膜血型糖蛋白A构象变化研究

为进一步验证麦胚凝集素作用下膜血型糖蛋白Ａ空间构象的变化,本文用傅里叶变换红外技术,定量测定了麦胚凝集素作用下,溶液血型糖蛋白Ａ和脂蛋白体膜血型糖蛋白Ａ的二级结构的改变,发现麦胚凝集素结合于血型糖蛋白Ａ导致血型糖蛋白Ａα—螺旋减少,β—结构增加,麦胚凝集素的抑制剂Ｎ—乙酰葡萄糖胺对麦胚凝集素诱导的血型糖蛋白Ａ二级结构的改变有抑制作用。本文同时用扫描隧道显微术直接观察了麦胚凝集素结合前后膜血型糖蛋白Ａ分子形态的变化。     

麦胚凝集素作用下膜血型糖蛋白A构象变化研究

为进一步验证麦胚凝集素作用下膜血型糖蛋白Ａ空间构象的变化,本文用傅里叶变换红外技术,定量测定了麦胚凝集素作用下,溶液血型糖蛋白Ａ和脂蛋白体膜血型糖蛋白Ａ的二级结构的改变,发现麦胚凝集素结合于血型糖蛋白Ａ导致血型糖蛋白Ａα—螺旋减少,β—结构增加,麦胚凝集素的抑制剂Ｎ—乙酰葡萄糖胺对麦胚凝集素诱导的血型糖蛋白Ａ二级结构的改变有抑制作用。本文同时用扫描隧道显微术直接观察了麦胚凝集素结合前后膜血型糖蛋白Ａ分子形态的变化。     

鸡胚肝乳糖凝集素专一性的研究——SEM-ARG技术的应用

本文利用扫描电镜放射自显影(SEM-ARG)技术研究鸡胚肝凝集素的专一性。该凝集素经乳糖尿素液抽提和离心分离后,再用DE-52纤维素柱和蓝色葡聚糖柱进一步纯化。纯化后的鸡胚肝凝集素用 ¹²⁵Ⅰ标记。以标记的 ¹²⁵Ⅰ-凝集素为探针再标记来自不同组织的细胞。标记的细胞经过放射自显影,用扫描电镜对细胞表面凝集素受体的位点进行直接观察。实验结果表明鸡胚肝凝集素对细胞的凝集作用具有相对的专一性。     

凝集素与细胞凝集 Ⅰ.伴刀豆球蛋白A及植物血凝素对正常细胞、胚胎及肿瘤细胞的凝集效应

自从 J.C.Aub 于1963年报道麦胚凝集素(WGA)对正常及肿瘤细胞的凝集作用有显著差异之后,已在数十种肿瘤及病毒转化的细胞也得到类似的结果。近年来又发现胚胎及分裂细胞也像肿瘤细咆一样易被凝集素凝集。因此,对凝集素的凝集效应与细胞癌变、繁殖和分化的关系,受到高度重视。     

鸡胚肝凝集素受体的放射自显影和扫描电镜观察

近些年来用标记过的外源凝集素作探针以研究细胞膜的结构和功能已取得很大进展。本文首次用~(125)I标记鸡胚肝凝集素,再以~(125)I-凝集素亲和标记鸡胚肝细胞。标记的鸡胚肝细胞经放射自显影(ARG),可用扫描电镜(SEM)对细胞表面受体的位点进行直接观察,以研究鸡胚肝凝集素受体的分布特征。     

水稻胚胎发育与萌发过程中凝集素活性、含量与免疫学性质的变化

稻胚凝集素(RGL)存在胚中,胚乳中没有测得凝集素活性。水稻开花后7～21天胚中RGL括性与含量迅速增加、积累,基本达到成熟胚的最高水平。在开花后7与13天胚中除了有RGL存在外还发现有与RGL免疫学性质无关的凝集素存在。在萌发早期RGL活性与含量迅速下降,在浸种萌发后1～4天之间则又保持相对恒定。水稻胚胎发育与萌发过程中没有观察到与RGL免疫学性质相关但分子性质不同的凝集素存在。RGL是稻胚发育过程中形成的专一蛋白质,它的表达与积累有严格的时空专一性,它的活性与含量变化与细胞分裂、分化等胚胎发育过程是相关联的。     

凝集素抑制林蛙卵裂沟新膜外露的作用

剥去受精膜的林蛙卵分裂时,分裂沟中的新膜会暴露出来。林蛙老膜上有大量均匀分布的麦胚和大豆凝集素受体。卵裂前,将剥去受精膜的蛙卵浸于上述的凝集素溶液中,新膜的外露就被抑制。凝集素愈浓,浸泡时间愈长,抑制愈大。在这些卵的表面可看到一层较厚的由凝集素引起的外被。碱处理过的受精卵表面,凝集素受体减少,凝集素抑制新膜外露的作用亦减弱,由凝集素引起的外被亦薄。凝集素是多价的,会在细胞表面产生交链,形成“外骨骼”,抑制新膜外露。凝集素也可通过受体,影响微丝,产生作用。     

稻胚中几种植物凝集素的内源受体

用4种不同的植物凝集素制成亲和吸附剂从成熟的稻胚中分离相应内源受体,发现“双丰1号”成熟稻胚中不合与麦胚凝集素(WGA)结合的物质,但含有少量与自身凝集素(S-RGL)和“寒丰”稻胚凝集素(K-RGL)结合的物质(0.1～0.2 mg/g胚),并含有较大量的与伴刀豆球蛋白(conA)结合的物质(1.0～1.5 mg/g胚)。用凝胶电泳分析受体组成,在低pH-不连续PAGE图谱中,conA受体有2条带,S-KGL和H-RGL受体均只有1条迁移率相同的带。SDS-IPAGE图谱显示,conA受体有7条多肽,S-RGL和H-RGL受体具有迁移率相同的8条多肽。说明两种受体的分子性质相同。     

农杆菌介导的半夏凝集素基因(Pinellia Ternate Agglutinin Gene,pta)对小麦的遗传转化及鉴定

以甘肃主要推广春小麦品种陇春22幼胚为转基因受体材料,建立了农杆菌介导的小麦遗传转化体系。以预培养4天的幼胚愈伤组织为受体,C58c1农杆菌菌株为供体,将含有半夏凝集素基因的重组质粒pBIpta转入了小麦,经G418 25 mg/L抗性筛选、PCR检测和荧光定量PCR检测共获得转基因植株3株,外源基因的插入拷贝数分别为2、1、3。同时对转基因小麦的T₁代植株进行了PCR检测和抗虫性分析,表明半夏凝集素基因在转基因植株的后代中得到了遗传并有一定的抗蚜虫作用。     

植物凝集素的超级家族

凝集素是一类专一、可逆地和糖类结合的蛋白质,迄今已经分离纯化并测定了氨基酸序列的凝集素已有不少,一些凝集素以及它们与配体糖相互结合的复合物的高级结构也已经给出,许多工作已深入到基因水平．就目前已有的知识,说明植物凝集素是一个庞大的蛋白质家族．     

Characterization of glycoside residues of porcine zona pellucida and ooplasm during follicular development and atresia

The glycoside residues (glycoconjugates, GC) of the zona pellucida (ZP) glycoproteins are important during the first phases of fecundation. Our aim in this work was to determine the lectin affinity pattern of porcine ZP in order to analyze the changes that take place during: (a) preantral folliculogenesis, (b) the follicular atresia process, and (c) antral growth. Several prepubertal and adult pig ovaries and different sized antral follicles were used. Conventional carbohydrate histochemical techniques and peroxidase and digoxigenin (DIG) lectins were used to reveal the acid groups and the glycosidic residues of the ZP. It was seen that the ZP forms in the preantral follicles throughout their growth period. In primordial and primary follicles, ZP in the process of formation showed neutral GC. SBA, RCA-I, MAA, WGA lectins, and AAA after methylation-saponification (MS) were positive in the ZP of primordial and primary follicles. The affinity for SBA, RCA-I, MAA, and WGA increased in the multilaminar-primary follicles and new affinities for UEA-I and LFA were observed. After MS, AAA, SNA, PNA, and SBA reactivity was observed. The ZP of antral follicle oocytes of different sizes showed the same lectin pattern as multilaminar-primary follicles. The oocyte ooplasm and the follicular fluid of large antral follicles showed less affinity for WGA and LFA lectins and less intensive staining with AB (pH 2.5). Atresia did not change the antral or preantral follicle oocyte ZP lectin pattern. In conclusion, the follicles showed substantial changes in their ZP glycosidic composition as they developed, especially, during the change from primary to multilaminar-primary follicles. The ZP glycosidic composition showed no significant change during the growth of antral follicles and follicular atresia in our study.     

Different effects of lectins on the ligand binding of the NMDA receptors and sigma sites in rat brain hippocampus synaptic membranes

The effects of the lectins concanavalin A, WGA, ricin, abrin, and the mistletoe lectins from Viscum album MLI, MLII, and MLIII on the binding of ligands of the NMDA and sigma receptors in rat hippocampus synaptic plasma membranes were investigated. Binding of [³H]MK-801, [³H]glutamate, [³H]5,7-DCKA, and [³H]glycine to the membranes was decreased by 40-60% after addition of galactose-specific lectins (mistletoe lectins MLI, MLII, ricin, abrin) at concentrations of 0.01 mg/ml, but was not affected by the glucose- and mannose-specific lectin Con A, an acetylglucosamine-specific lectin WGA, or an acetylgalactosamine-specific lectin MLIII. The binding of [³H]SKF 10047 was decreased only in the presence of MLIII and did not change after addition of the other lectins. It is suggested that lectin-sensitive ligand binding sites of sigma- and NMDA receptors are located separately, and that the carbohydrate side chains of the sigma receptor do not participate in the modulation of the NMDA-receptor.     

Novel effect of plant lectins on the inhibition of Streptococcus mutans biofilm formation on saliva-coated surface

Aims:  Dental caries is caused by the disturbance in oral homeostasis, marked by a notable increase in the population of Streptococcus mutans . Lectins are a group of plant proteins that are capable of recognizing the glycoconjugates present on the bacterial surface. The aim of this study was to evaluate the effect of seven plant lectins on the growth and initial adhesion of S. mutans .
Methods and Results:  Lectins of different carbohydrate specificities were isolated from plant sources by conventional methods of protein purification. The effect on growth of S. mutans was evaluated following CLSI guidelines. None of the lectins used in this study inhibited the bacterial growth and multiplication. The adherence and biofilm formation of bacteria to saliva-coated polystyrene plates was tested in the presence of plant lectins. All the plant lectins tested, inhibited both the adherence and biofilm in a concentration dependent manner. Confocal microscopy and scanning electron microscopy were employed to assess the biofilm formation in the presence of plant lectin (glucose/mannose-specific) at sub-minimal inhibitory concentrations. These evaluations revealed that lectins inhibited the clumping and attachment of S. mutans .
Conclusions:  Lectins tested here inhibited initial biofilm formation by S. mutans. Glucose/Mannose-specific lectin altered the adhesion arrangement of the bacteria on the saliva-coated surfaces.
Significance and Impact of the Study:  The plant lectins used in this study may offer a novel strategy to reduce development of dental caries by inhibiting the initial adhesion and subsequent biofilm formation of S. mutans.     

Lectin Analysis of Surface Saccharides in Two Trichomonas vaginalis Strains Differing in Pathogenicity*

SYNOPSIS. Surface saccharides in 2 Trichomonas vaginalis strains, the moderately pathogenic, JH34A, and the mild, JH162A, were analyzed with the aid of plant lectins. Concanavalin A (Con A), wheat germ agglutinin (WGA), soybean agglutinin (SBA), castor bean agglutinin (CBA), and lectin from the garden pea (GPA) were employed in agglutination tests and in treatment of ultrathin sections for electron microscopy according to the horseradish peroxidase-3,3′-diaminobenzidine method. With Con A and WGA, small quantitative differences were noted between the 2 strains in the results of agglutination and in the reaction-product deposits observed by electron microscopy. Distribution of the binding sites for the 2 lectins was also somewhat different in the JH34A and JH162A trichomonads. In general, the reactions with the more pathogenic strain were slightly stronger. Although the reactions with SBA and CBA lectins were weaker than those with Con A or WGA, they provided the means for qualitative differentiation between the 2 trichomonad strains. SBA alone agglutinated the JH34A strain and formed demonstrable deposits on the cell surfaces. On the other hand, only CBA reacted with JH162A flagellates. The garden pea lectin failed to bind to the surface of either strain. On the basis of results obtained with the control preparations incubated in the presence of specific inhibitors, it was concluded that both strains had α-methyl-D-mannoside and/or α-methyl-D-mannoside-like as well as N-acetyl-D-glucosamine residues on their surfaces. In addition, JH34A strain had D-lactose-containing residues while JH162A trichomonads had residues with D-galactose. Neither strain appeared to possess residues containing N-acetyl-D-galactosamine.     

Lectin-mediated sperm agglutination of bufo arenarum and leptodactylus chaquensis spermatozoa

Various plant lecins were employed in cell agglutination experiments to ascertain the presence of specific saccharides in the surface of B arenarum and L chaquensis spermatozoa. B arenarum spermatozoa were specifically agglutinated with Concanavalin A (Con A), phytohemagglutinin P (PHA-P), and wheat germ agglutinin (WGA), but not with soybean agglutinin (SBA). In contrast, L chaquensis spermatozoa were strongly agglutinated by SBA, WGA, and PHA-P. L chaquensis spermatozoa did not agglutinate with Con A even at high concentrations. Lectinmediated sperm agglutination was inhibited in the presence of specific lectinbinding sugars. Spermatozoa from both species were agglutinated randomly with all lectins suggesting a uniform distribution in the sperm surface of the lectinbinding saccharide ligands. B arenarum sperm agglutination induced by Con A is sensitive to temperature. B arenarum spermatozoa are more agglutinable at 24°C than at 4°C. These results suggest that lectin-binding site mobility is necessary for sperm agglutination.     

Effects of wheat germ agglutinin on tunicate egg activation and fertilization: Is there a plasma membrane sperm receptor system on Ascidia ceratodes eggs?

Little work has been carried out on the sperm recognition systems present on the egg plasma membrane. Here it is shown that wheat germ agglutinin (WGA) interferes with the sperm-interacting system on the plasma membrane of eggs of the ascidian, Ascidia ceratodes. The WGA activates the dechorionated egg, indicating that a plasma membrane sugar residue can be directly tied to egg activation. Low concentrations of this lectin do not activate the eggs, but reduce fertilizability. This observation suggests that the WGA binding site might be part of a sperm reception–activation complex in the plasma membrane. While WGA also affects sperm binding to the chorion, the mechanisms of sperm interaction at the plasma membrane and chorion show different sensitivities to lectins, sugars and enzymes.     

Lectinomics I. Relevance of exogenous plant lectins in biomedical diagnostics

This review focuses on utilization of plant lectins as medical diagnostic reagents and tools. The lectin-related diagnostic is aimed at detection of several diseases connected to alteration of the glycosylation profiles of cells and at identification of microbial and viral agents in clinical microbiology. Certain lectins, proposed for or used as diagnostic tools could even recognize those cellular determinants, which are not detected by available antibodies. Broad information is presented on the lectinomics field, illustrating that lectin diagnostics might become practical alternative to antibody-based diagnostic products. In addition, the rising trend of lectin utilization in biomedical diagnostics might initiate a development of innovative methods based on better analytical technologies. Lectin microarray, a rapid and simple methodology, can be viewed as an example for such initiative. This technology could provide simple and efficient screening tools for analysis of glycosylation patterns in biological samples (cellular extracts, tissues and the whole cells), allowing thus personalized detection of changes associated with carbohydrate-related diseases.     

Alterations in sugar residues in squamous metaplasia in hamster tracheal explants induced by benzo(a)pyrene and its reversal by retinoic ACID

Summary We have demonstrated that squamous metaplasia induced by benzo(a)pyrene (BP) in the hamster tracheal explants accompany distinct alterations in carbohydrate moieties in the epithelial mucosa. Most prominent alterations were the preferential binding of peanut agglutinin (PNA) and wheat germ agglutinin (WGA) in the basal cell layer in metaplastic lesions. In this study we examined if reversal of BP-induced lesions by all-trans retinoic acid (RA) results in the acquisition of normal carbohydrate composition by the tissue. Four lectins, PNA, WGA, Dolichos biflorus agglutinin, and Concanavalin A, in their horseradish peroxidase conjugates were used. In control explants the intercellular plasma membrane of basal and mucous cells exhibited no significant reaction with any of the lectins tested. In the metaplastic lesions induced by BP, PNA and WGA intensely stained the plasma membrane and intercellular spaces of basal and intermediate cell layers; the granular layer cells did not bind PNA whereas they were stained moderately with WGA. RA, which reversed the metaplasia, also conferred the tissue with lectin binding patterns similar to that of control explants. These results thus show that the reversal of metaplasia is accompanied by acquisition of the tissue’s original carbohydrate composition. This research was supported by grant RO1-HL32308 from the National Heart, Lung and Blood Institute, Bethesda, MD.     

Cell Surface Saccharides in Three Phytomonas Species Differing in Host Specificity

ABSTRACT. Cell surface carbohydrates of three phytoflagellates, Phytomonas francai. Phytomonas serpens and Phytomonas sp. from different hosts including cassava, coreid insect Phthia picta and the milkweed plant Euphorbia hyssopifolia, respectively, were analysed by agglutination assays employing a battery of highly purified lectins with affinity for receptor molecules containing N-acetylglucosamine (d-GlcNAc), N-acetylgalactosamine (D-GalNAc), galactose, mannose-like (D-Man-like) residues and fucose, and by binding assay using radiolabeled [¹²⁵I]-wheat germ agglutinin (WGA) and fluorescent WGA lectin, as well as glycosidases of known sugar specificity, Escherichia coli K with mannose-affinity fimbrial lectin was also used as an agglutination probe. In general, the presence of D-GlcNAc. D-GalNAc and D-Man-like residues was detected in the phytomonads' plasma membrane. These sugar moieties were confirmed in whole cell hydrolysates as assessed by gas-liquid chromatography (GLC) which in addition, also showed the presence of galactose and xylose. However, marked differences in cell surface carbohydrate structures were observed. Wheat germ agglutinin, which binds to sialic acid and/or d-GlcNAc-containing residues, shows selective agglutinin activities for P. francai and Phytomonas sp., while Bandeiraea simplicifolia II agglutinin (which recognizes d-GlcNAc units) specifically bound to Phytomonas sp. Helix pomatia agglutinin which binds to D-GalNAc-containing residues reacted preferentially with Phytomonas sp. and P. serpens. Con A, which recognizes D-Man-like receptors, agglutinates all the phytomonads; however, the higher interaction was observed with Phytomonas sp. P. francai was selectively agglutinated in the presence of E. coli fimbrial lectin. Fluorescence WGA binding was significantly decreased by N-acetylglucosaminidase activities and the cell agglutination was not altered by neuraminidase treatment, suggesting the presence of an exposed D-GlcNAc moiety on the P. francai and Phytomonas sp. surfaces. Binding studies with [¹²⁵I]-WGA essentially confirmed the fluorescence WGA binding and agglutination assays.