Purification and separation of tomato isolectins by chromatofocusing

Tomato lectin can be rapidly separated from a crude extract of tomato fruit proteins by chromatofocusing. The lectin is recovered from a PBE94 column in two peaks, each with a specific activity comparable to that of lectin purified by affinity chromatography on ovomucoid-Sepharose. Both isolectins consist of a single polypeptide chain (Mr 68,000) and have similar properties.     

Crystal structures of carbohydrate recognition domain of blood dendritic cell antigen‐2 (BDCA2) reveal a common domain‐swapped dimer

We report on crystal structures of a carbohydrate recognition domain (CRD) of human C‐type lectin receptor blood dendritic cell antigen‐2 (BDCA2). Three different crystal forms were obtained at 1.8–2.3 Å resolution. In all three, the CRD has a basic C‐type lectin fold, but a long loop extends away from the core domain to form a domain‐swapped dimer. The structures of the dimers from the three different crystal forms superimpose well, indicating that domain swapping and dimer formation are energetically stable. The structure of the dimer is compared with other domain‐swapped proteins, and a possible regulation mechanism of BDCA2 is discussed. Proteins 2014; 82:1512–1518. © 2013 Wiley Periodicals, Inc.     

Mannose-Binding Lectin Serum Levels are Low in Persons with Clinically Active Coccidioidomycosis

Background  Mannose-binding lectin (MBL) is a circulating collectin that is part of the innate immune response. We explored the serum levels of MBL in persons with different forms of coccidioidomycosis. Methods  Serum MBL was measured by ELISA from samples obtained from healthy donors with immunity to Coccidioides, and those with various forms of active coccidioidomycosis. Blood cell specimens from a subgroup of subjects with active coccidioidomycosis were examined for single nucleotide polymorphisms of the MBL gene and promoter regions. Results  The control group comprised 29 healthy immune subjects. Patient groups with active coccidioidomycosis consisted of 20 patients with symptomatic primary pulmonary coccidioidomycosis, 26 with non-meningeal disseminated coccidioidomycosis, and nine with coccidioidal meningitis. The group with active coccidioidomycosis was significantly older and more likely to be male than the control group (for both, P < 0.001). The mean ± SEM level of serum MBL in the healthy controls was 169.4 ± 28.6 ng/ml, significantly higher than the 79.2 ± 10.9 ng/ml for all active groups (P < 0.001). Moreover, the active coccidioidomycosis group was significantly more likely to have serum MBL level ≤70 ng/ml compared to the control group (P = 0.001). Genetic analysis in 27 subjects with active coccidioidomycosis revealed marked variation based on race and ethnicity. Among a subgroup of 10 white, non-hispanic men with active coccidioidomycosis, there was a significant association between the H and P haplotypes and MBL levels ≤70 ng/ml (P < 0.036 and P < 0.035, respectively). Conclusions  These data suggest that there is an association between low serum MBL levels and symptomatic coccidioidomycosis.     

酵母多糖影响人朗格汉斯细胞C型凝集素受体的研究

探究酵母多糖（Zymosan）对人朗格汉斯细胞（Langerhans cells, LCs）C型凝集素受体（C-type lectin receptors, CLRs）的影响。收集健康志愿者血液样本,Ficoll淋巴细胞分离液分离外周血单个核细胞,CD14磁珠分选CD14⁺细胞,用含有GM-CSF（1 000 U/mL）、IL-4（1 000 U/mL）、TGFβ-1（10 ng/mL）的培养液诱导培养LCs, Zymosan刺激LCs,8、24 h后收集细胞。提取Zymosan实验组和对照组细胞RNA,实时RT PCR分析5种CLRs（Dectin-1、Dectin-2、DC-SIGN、Mincle、MRC-2） mRNA的水平,扩增产物测序鉴定。共检测10个个体来源LCs在酵母多糖刺激后其CLRs mRNA变化的情况。在10组样本中,有6组Dectin 2受体mRNA较对照组增加（变化倍数≥2）,其中3组增加明显（变化倍数≥5）,最大增加倍数为20.91;有5组Mincle受体mRNA较对照组增加（变化倍数≥2）,其中4组增加明显（变化倍数≥5）,最大增加倍数为19.98;有1组MRC-2受体mRNA较对照组明显增加（变化倍数≥5）。Dectin-1和DC-SIGN受体mRNA没有增加。Zymosan能增加部分个体来源LCs内Dectin-2、Mincle和MRC-2受体mRNA的水平,而对Dectin-1和DC-SIGN受体mRNA的水平没有影响。本研究结果完善了Zymosan调节机体免疫功能的作用机制。     

Purification and characterization of Kurloff cell sialoglycoproteins with acid phosphatase activity

The majora2–6 sialoglycoproteins in detergent-extracts of Kurloff cells were purified by anion-exchange andSambucus nigra agglutinin-affinity chromatographies. The similar ultrastructural localisations of (1)S. nigra agglutinin-gold conjugates and (2) acid phosphatase activities on the Kurloff body and particularly on its myelin figures indicated that the majora2-6 sialoglycoproteins of the Kurloff cell had acid phosphatase activity. Two-dimensional electrophoresis showed that these tartrate-sensitive phosphatases corresponded to 2 acidic (pI 3.4–3.7) polypeptides of 36 and 34 kDa. Hydrolysis with peptide-N-glycosidases F gave a 33 kDa apoprotein rich in alanine, glutamic acid, tyrosine and lysin. A lectin-affinity study demonstrated that they contained hybrid type bisected and fucosylatedN-linked oligosaccharides. Cytotoxic properties were previously attributed to Kurloff cells and other studies suggested that not only acid phosphatases but alsoa2-6-linked sialic acid residues themselves may participate in natural killer activity.     

Localization of wheat-germ agglutinin in developing wheat embryos and those cultured in abscisic acid

The time course of appearance of wheat-germ agglutinin (WGA) in the various embryonic tissues during embryogenesis in Triticum aestivum L. was studied by sensitive immunofluorescence and peroxidase-antiperoxidase detection systems. The radicle, root cap and coleorhiza first accumulated WGA in early Stage II (8-10 d post-anthesis) prior to the main period of embryo growth, while WGA was found in the epiblast and coleoptile in early and late State III, respectively. Stage III is characterized by maximum embryo growth, followed by desiccation which occurs in Stage IV. When Stage-II embryos were precociously germinated in the absence of abscisic acid (ABA) no WGA was detected in the coleoptile and epiblast of the young seedlings. In the presence of ABA, Stage-II embryos did not germinate but WGA precociously accumulated in the coleoptile and epiblast. The levels and distribution of WGA in the resulting embryo resembled those in a fully mature, dry embryo (Stage V). Barley possesses a seed lectin similar to WGA, but it is never detected in coleoptiles. Some but not all of the barley cultivars tested were found to accumulate lectin in this organ of mature embryos when treated with ABA. Thus, ABA appears to be involved in the highly regulated temporal and spatial expression of WGA during embryogenesis in cereals.Abbreviations ABA abscisic acid - DIC differential interference contrast - PAP peroxidase-antiperoxidase - WGA wheat-germ agglutinin     

Lectin in five soybean cultivars previously considered to be lectin-negative

Hemagglutinating proteins were isolated by affinity chromatography from seeds of each of five cultivars of soybeans (Clycine max (L.) Merr.) previously reported to lack detectable lectin (S.P. Pull et al., 1978; Science 200, 1277). Quantities were between 1,000 and 10,000 times less than that found in the seeds of the reference cultivar, Chippewa. The sensitivity of the hemagglutinating assay was 0.05 g ml^-1. Hemagglutinating activity was demonstrated in affinity-purified fractions from bulk seeds and seeds from individual plants in two cultivars, 30–70% ammonium-sulfate-precipitable fractions of seeds from individual plants of all five cultivars, and in whole crude extracts of individual seeds from each cultivar. In all instances, hemagglutinating activity was inhibited by galactose, anti-soybean agglutinin (SBA), and lectin-binding polysaccharide produced by Rhizobium japonicum. Affinity-purified lectin from seeds of a single Columbia plant was labeled with fluorescein isothiocyanate (FITC) and observed by fluorescence microscopy to bind to R. japonicum cells with specificity, intensity and localization indistinguishable from FITC-SBA. Lectins from distinguishable from FITC-SBA. Lectins from three cultivars in sufficiently high concentration for study had molecular properties very similar to Chippewa SBA.Abbreviations FITC fluorescein isothiocyanate - IgG immunoglobulin G - SBA soybean agglutinin