HEp-2细胞MPM-2磷蛋白的蛋白质组学分析

MPM-2磷蛋白家族成员在有丝分裂进程中起着重要作用．为了从MPM-2磷蛋白家族层面入手研究细胞周期调控中磷蛋白的重要作用，以人上皮样喉癌细胞株HEp-2为细胞模型，运用双向电泳、免疫印迹以及质谱技术等蛋白质组学的方法对其细胞抽提物中的部分MPM-2磷蛋白进行了分析鉴定，初步确定了乙二醛酶(GlyoxalaseⅠ)、真核翻译起始因子(eIF4B)以及核质蛋白NPM等MPM-2特异识别的细胞磷蛋白，这将有助于进一步研究MPM2磷蛋白家族成员在细胞周期调控中的重要作用．     

Affinity enrichment of plasma membrane for proteomics analysis

Proteomics analysis of plasma membranes from cells exposed to different extracellular environments is potentially a powerful approach for the identification of membrane-associated proteins responding to these environments. Preparation of high concentration plasma membrane fractions with low contamination from cellular organelles is essential for such studies. Here, we describe an affinity enrichment method, which combines cell surface biotinylation with affinity enrichment by immobilized streptavidin beads, for the isolation of plasma membranes. This method results in a 400-fold enrichment of plasma membrane relative to endoplasmic reticulum, a major contaminant in standard plasma membrane preparations, and dramatically reduces contamination from other cellular organelles. The biotinylation reaction did not interfere with ligand-dependent activation of receptor tyrosine kinases or G-protein coupled receptors, suggesting cell-surface signal transduction machinery remains functional. Membrane fractions prepared by this method should provide excellent starting materials for membrane proteomics analysis such as studies of dynamic trafficking and regulation of signaling molecules or identification of disease-specific membrane markers.     

Relative Abundance of Alpha-Amylase/Trypsin Inhibitors in Selected Sorghum Cultivars

Sorghum is of growing interest and considered as a safe food for wheat related disorders. Besides the gluten, α-amylase/trypsin-inhibitors (ATIs) have been identified as probable candidates for these disorders. Several studies focused on wheat-ATIs although there is still a lack of data referring to the relative abundance of sorghum-ATIs. The objective of this work was therefore to contribute to the characterization of sorghum ATI profiles by targeted proteomics tools. Fifteen sorghum cultivars from different regions were investigated with raw proteins ranging from 7.9 to 17.0 g/100 g. Ammonium bicarbonate buffer in combination with urea was applied for protein extraction, with concentration from 0.588 ± 0.047 to 4.140 ± 0.066 mg/mL. Corresponding electrophoresis data showed different protein profiles. UniProtKB data base research reveals two sorghum ATIs, {"type":"entrez-protein","attrs":{"text":"P81367","term_id":"3913899","term_text":"P81367"}}P81367 and {"type":"entrez-protein","attrs":{"text":"P81368","term_id":"3913900","term_text":"P81368"}}P81368; both reviewed and a targeted LC–MS/MS method was developed to analyze these. Quantifier peptides ELAAVPSR ({"type":"entrez-protein","attrs":{"text":"P81367","term_id":"3913899","term_text":"P81367"}}P81367) and TYMVR ({"type":"entrez-protein","attrs":{"text":"P81368","term_id":"3913900","term_text":"P81368"}}P81368) were identified and retained as biomarkers for relative quantification. Different reducing and alkylating agents were assessed and combination of tris (2 carboxyethyl) phosphine/iodoacetamide gave the best response. Linearity was demonstrated for the quantifier peptides with standard recovery between 92.2 and 107.6%. Nine sorghum cultivars presented up to 60 times lower ATI contents as compared to wheat samples. This data suggests that sorghum can effectively be considered as a good alternative to wheat.     

蛋白质组学技术在动物科学中的研究进展

蛋白质组学在后基因组时代以其独特的角度和重要的作用已成为生命科学领域研究的热点内容。动物科学作为生命科学的一个重要分支,其相关研究对人类的生活有着重要意义。为了系统地了解和展望蛋白质组学技术在动物科学中的研究进展,综述了蛋白质组学的概念及其相关技术,归纳和评析了近年来蛋白质组学在动物繁殖、营养、遗传育种和疾病防治方面的研究现状。     

XapR蛋白在嗜水气单胞菌抗生素耐药性的功能

研究嗜水气单胞菌（Aeromonas hydrophila） LysR型转录调节因子中XapR蛋白抗生素耐药的调控机制。通过测定嗜水气单胞菌ATCC 7966（野生型菌株）与嗜水气单胞菌ATCC 7966 xapR基因缺失株（ΔxapR）对34种抗生素的最低抑菌浓度,系统地评价xapR基因抗生素耐药性;进一步利用定量蛋白质组学技术,探讨xapR参与嗜水气单胞菌调控的生物过程。结果表明：xapR基因缺失后细菌对头孢孟多的耐药性减弱;xapR基因可调控细菌碳代谢、TCA循环等多种代谢途径,此外还与外膜蛋白、TonB、ABC转运系统等抗性基因的表达密切相关。综上,XapR蛋白可通过调节细菌中心代谢途径以及多个抗性基因改变嗜水气单胞菌的耐药性。     

Profiling and comprehensive expression analysis of ABC transporter solute-binding proteins of Bacillus subtilis membrane based on a proteomic approach

We analyzed ABC transporter solute-binding proteins (SBPs) of the Bacillus subtilis membrane using a proteomic approach. We prepared a washed cell membrane fraction that was insoluble in 134 mM nondetergent sulfobetaine and then extracted proteins using mixtures of detergents in a stepwise manner. The membrane proteins were resolved by three two-dimensional polyacrylamide gel electrophoresis (2-D PAGE) or two one-dimensional (1-D) PAGE procedures, electroblotted, and digested in the presence of 5% or 80% acetonitrile. Thereafter, matrix-assisted laser desorption/ionization-time of flight-mass spectrometry (MALDI-TOF MS) identified 637 proteins corresponding to 15.9% of the total cellular proteins. We predicted that among these, 256 were membrane proteins, 101 were lipoproteins or secretory proteins and 280 were soluble proteins containing peripheral proteins that function in both the cytoplasm and the cell membrane such as SecA and FtsY. Among the 637 proteins, we identified 30 SBPs among 38 importers predicted by a bioinformatic search of the genome. We confirmed expression of the genes for the 30 SBPs using DNA microarray analysis. We compared the 2-D gel separation profiles of submembrane fractions solubilized by 1% n-dodecyl-beta-D-maltoside from cells cultured on Luria Bertani (LB), S7, and S7 medium without glutamate as well as DNA microarray data on LB and S7. The results suggested that YcdH, YtmK and YurO are binding proteins for Mn(++), glutamate and glucose, respectively, and that YqiX and YxeM are binding proteins for amino acids (tryptophan in S7 medium).     

蛋白质组学在人乳蛋白质研究中的应用

人乳是新生儿最理想的天然食物，蛋白质是人乳中最主要的营养成分之一。随着蛋白质组学技术的发展，利用蛋白质组学的方法研究人乳蛋白质也取得了一些研究成果。本文综述了近年来蛋白质组学技术在人乳蛋白质研究中的应用，分别从人乳蛋白质的组成研究、动态变化、人乳与其他来源乳汁的蛋白质差异比较、人乳磷酸化蛋白和糖基化蛋白研究、人乳内源肽的研究及人乳蛋白与疾病等几个方面进行了阐述。蛋白质组学技术使人乳蛋白质的研究进入了微量营养研究的时代，人乳蛋白质组学的研究成果将为母婴健康提供更好的保障。     

蛋白质组学和代谢组学在微生物代谢工程中的应用

构建微生物细胞工厂是化学品、生物能源以及药物分子可持续生产的可行性策略。然而，微生物的代谢复杂、调控严谨，制约着目标产物高效合成。蛋白质组学和代谢组学可以从系统生物学角度分析酶和代谢物组分，从而理解复杂的生物系统，为微生物代谢工程改造提供重要线索。该文介绍了蛋白质组学和代谢组学在微生物代谢工程中的应用，包括基因组尺度代谢模型构建、菌株生物合成优化、指导菌株耐受性改造、限速步骤预测、植物次级代谢途径挖掘，从而为微生物合成天然产物提供新的基因或途径。在此基础上，该文还展望了生物大数据未来的发展方向。     

蛋白质与蛋白质组学的研究进展

介绍了蛋白质、蛋白质组和蛋白质组学的基本概念，蛋白质组学的研究内容、方法技术、形成与发展。