免疫蛋白质组学及疫苗靶位筛选

蛋白质组学自建立起即向生命科学的其他研究领域渗透,形成了多样的交叉学科。免疫蛋白质组学即由蛋白质组学与蛋白质免疫印迹技术相结合而产生的一个新研究方向,在免疫原性蛋白质研究及疫苗靶位筛选中展示出广泛的应用前景。该文就免疫蛋白质组学的形成、主要研究技术体系及其进展、在免疫原性蛋白质鉴定、新型高效候选疫苗靶位发现等方面进行概述。973计划(No.2006CB101807)资助项

Studying the growth metabolism of Aeromonas hydrophila with the methods of proteomics

采用蛋白质组学的方法之一二维电泳技术 ,对嗜水气单胞菌生长周期中各不同阶段的蛋白质组成进行了分析。结果显示 ,在不同生长阶段 ,其蛋白质组成均发生明显变化。发现了与细菌生长代谢各阶段生理活动有关的差异蛋白质 ,有助于深入了解嗜水气单胞菌生长周期中不同阶段的基因组表达情况。同时 ,还对二维电泳和普通SDS 聚丙烯酰胺凝胶电泳进行比较。结果表明 ,二维电泳技术是研究蛋白质组学的有效工具Aeromonas hydrophila is one of the important pathogenic bacteria of aquaculture. It is significant, therefore, to study the changes on functional genome in its growth cycle. Two dimensional electrophoresis (2 D PAGE), one of the approaches for proteomics, was used to study protein composition in different phases during the cycle. The result showed that the composition was significantly changed in different phases. The discovery of different pertinent proteins in the different growth phases will contribute to find out the genome expression of A. hydrophila in different growth phases. In this experiment 2 D PAGE was compared with normal SDS PAGE in the same conditions, and the result indicated that 2 D PAGE was an effective technique for proteomics study.国家自然科学基金资助项目 (3 9770 5 85

PRODUCTION OF iNOS INDUCED BY RICIN IN HUMAN HEPATOMA——STUDY OF IMMUNOHISTOCHEMISTRY

应用免疫组织化学方法观察不同浓度蓖麻毒素作用于肝癌细胞不同时间后 ,对 i NOS的诱导作用。结果显示 ,未受蓖麻毒素作用的肝癌细胞胞浆 i NOS呈阴性反应 ;当受到蓖麻毒素诱导后 ,才能在胞浆内合成。不同浓度的蓖麻毒素对肝癌细胞诱导 i NOS的产生没有明显差别。而蓖麻毒素诱导 i NOS的表达有明显的时间依赖性 ,2 h没有表达 ,4h才出现 ,随着时间的延长 ,到 8h时仍然有明显的表达。由于 i NOS的合成涉及基因转录、蛋白质合成等过程 ,故需诱导数小时后显示酶活性 ,但一经诱导生成 ,酶活性持续时间长。蓖麻毒素诱导 i NOS的产生在抗癌应用中具有重要的价值To study iNOS production in hepatoma stimulated by ricin at different concentration and at different time. The results demonstrated, iNOS is negative in cytoplasm when hepatoma was not stimulated by ricin. After stimulating hepatoma with ricin, iNOS is positive in cytoplasm. The iNOS productin has not obviously difference when stimulating hepatoma by ricin at different concentration. However, iNOS express of ricin induction has distinctly tome dependent from 2h to 8h. The iNOS activity has persisted for a number of hours in hepatome cells. Because iNOS synthesis has related to gene transcription, protein synthesis and etc. The iNOS production of ricin induction may contribute to the clinical application. Our results indicate that ricin has great potential for the treatment of hepatoma cells.福建省自然科学基金资助项目 (NO.C0 0 10 0 0 3);; 中国博士后科学基金资助项

Diagonal polyarcylamide gel electrophoresis in proteomics research

经典的蛋白质组学研究方法包括IEF/SDS-PAGE双向电泳和质谱技术的联用,但由于IEF的一些不足,限制了其应用范围。对角线电泳是蛋白质组学研究中的一项特殊分离技术,由于其原理与IEF/SDS-PAGE不同,正逐渐成为蛋白质组学中电泳分离技术的重要补充,特别是在膜蛋白和蛋白质相互关系的研究中将起到重要作用。本文综述了对角线双向电泳技术的特点、发展和在蛋白质组学研究中的最新进展,比较了双向电泳和对角线电泳的优缺点,展望了对角线电泳在蛋白质组学研究中的应用前景。The classical approach in proteomic analysis couples IEF/SDS-PAGE with mass spectrum.However,the shortcomings of IEF restrict its widespread applications.Diagonal polyarcylalmide gel electrophoresis,a special protein separation technique,shows its valuable application in proteomic research,especially in mem-brane proteins and protein-protein interaction because of its different feature characteristics from IEF/SDS-PAGE.This paper summarizes the characters,development and the latest applications of diagonal polyarcylamide gel electrophoresis in proteomics and the comparison between IEF/SDS-PAGE and the diagonal gel electrophoresis.国家“863”高科技发展计划项目(No.2004AA626030

微生物蛋白质组学研究进展

教育部优秀骨干教师基金资

Advancement on the function of hemocyanin

广东省自然科学基金(博士启动基金)资助项目(04300750);; 厦门市科技发展资助项

A Novel Cytoplasmic Protein with RNA-binding Motifs Is an Autoantigen in Human Hepatocellular Carcinoma

In hepatocellular carcinoma (HCC), autoantibodies to intracellular antigens are detected in 30–40% of patients. Patients with chronic hepatitis or liver cirrhosis develop HCC, and when this occurs, some patients exhibit autoantibodies of new specificities. It has been suggested that these novel autoantibody responses may be immune system reactions to proteins involved in transformation-associated cellular events. One HCC serum shown to contain antibodies to unidentified cellular antigens was used to immunoscreen a cDNA expression library, and a full length cDNA clone was isolated with an open reading frame encoding 556 amino acids with a predicted molecular mass of 62 kD. The 62-kD protein contained two types of RNA-binding motifs, the consensus sequence RNA–binding domain (CS-RBD) and four hnRNP K homology (KH) domains. This protein, provisionally called p62, has close identity (66–70%) to three other proteins at the amino acid sequence level, and all four proteins may belong to a family having CS-RBD in the NH2-terminal region and four KH domains in the mid-to-COOH– terminal region. The homologous proteins are: KH domain–containing protein overexpressed in cancer (Koc); zipcode binding protein, a protein which binds to a conserved nucleotide element in chicken β-actin mRNA (ZBP1); and a protein which binds to a promoter cis element in Xenopus laevis TFIIIA gene (B3). p62 protein is cytoplasmic in location, and autoantibodies were found in 21% of a cohort of HCC patients. Patients with chronic hepatitis and liver cirrhosis, conditions which are frequent precursors to HCC, were negative for these autoantibodies, suggesting that the immune response might be related to cellular events leading to transformation. However, the possible involvement of p62 autoantigen as a factor in the transformation process remains to be elucidated

Centralized Management To Share Resource

就"211工程"一期实验室建设管理经验进行总结,为二期建设提供参考。The article summarized the experience of lab construction and management in the first stage of Project 211, which offers reference for the construction in the second stage

IgA IMMUNE RESPONSES IN PATIENTS WITH HIV INFECTION

目的深入评价HIV感染者的IgA类免疫反应。方法将IgA类免疫反应产物分解为游离IgA、复合IgA和IgA/C3双特异性免疫复合物（IgA/C3—TCIC）三种组分，分别采用单向扩散法和ElISA技术进行分析。结果对68例无症状HIV感染者、120例AIdS患者和36例健康对照者的研究表明，HIV感染者的游离和复合IgA显著升高，IgA/C3—TCIC则明显下降，三者变化均与病情有一定关系，且其水平均在P24阳性组显著高于P24阴性组。游离IgA与Cd4和Cd4/Cd8比率呈负相关，而与复合IgA和IgA/C3—TCIC呈正相关，但复合IgA和IgA/C3—TCIC间无明显相关性。结论不同IgA类组分的免疫学意义有差异，其中复合IgA和IgA/C3—TCIC是判断HIV感染早期及其进展的良好指标，而游离IgA不失为反映HIV感染后期的一个特征性指标。患者血清IgA显著升高可能与抑制性T细胞的数量下降和功能障碍有关。并提出可考虑通过干扰复合IgA的形成和促进复合IgA的清除等途径来作为治疗疾病的新手段。Aim To Further understand IgA immune responses in patients with HIV inFection.Methods IgA immune responses were dissected into their components,namely Free IgA,complexed IgA and IgA/C3 two-component-determined circulating immune complexes (IgA/C3 -TCIC ), which were detected by single radio immune diFFusion or ELISA.Results Our study group included 36 HIV-seronegative controls, 68 asymptomaticHIV +subjects, 120 HIV + individuals with AIDS- related opportunistic inFection.Both levels of Free and complexed IgA were signiFicantly increased in patients,but the amount of IgA/C3-TCIC was markedly reduced among them.All of the three markers were dramatically higher in p24+group than p24-one.Free IgA was negatively associated with CD4 and CD4/CDS ratio,and positively with .complexed IgA and IgA/C3 -TCIC.However,there were no relationship between complexed IgA and IgA/C3-TCIC.Conclusion These Findings show thatcomplexed IgA and IgA/C3-TCIC may be better markers of early stage progression of HIV inFection,and thatFree IgA may be a more valuable indicator in later stage disease.The mechanism of elevated serum IgA may be involved in CD4 numbers and Function.Furthermore,a need For novel therapeutic strategies to modulate IgA immune responses in AIDS is suggested.加拿大国际发展总署;福建省自然科学基金!C9701

Comparative Proteomics of Proteins in Sinonovacula constricta Hemolymph under Low Salinity

应用双向聚丙烯酰胺凝胶电泳技术及图像分析技术寻找缢蛏在低盐环境压力下血淋巴内40kD以下小分子蛋白质组差异,制成肽质指纹图谱,MALDI-TOF-MS测序,通过数据库检索快速鉴定蛋白质或多肽。研究结果表明:低盐压力下,缢蛏血淋巴内有多种蛋白质发生变化,共鉴定出包括磷脂酶A2(Phospho lipaseA2)、锌指蛋白(ZincFingerProtein)等6种蛋白或多肽,其功能涉及营养、基因表达及蛋白合成调控等方面,说明其血淋巴在缢蛏的防御体系中具有关键性作用。The differences of the whole protein(under 40 kD) expression profiles in Sinonovacula constricta hemolymph after shot-term treatment of low salinity are detected using two-dimensional electrophoresis and image analysis software,The peptide mass map is obtained with MALDI-TOF-MS,A novel protein or polypeptide is identified by database retrieval.Several proteins or polypeptides in S. constricta hemolymph change under low salinity stress, including Phospholipase A2, Zinc finger protein and so on, which are of multiple functions such as nutrition, regulatory gene expression and protein synthesis, and so on. It′s obvious that hemolymph is critical in S. constricta defense mechanism.福建省教育厅资助项目(A类)(JA02179