蜂毒肽前体蛋白cDNA的克隆及序列分析

从蜜蜂（Ａｐｉｓｍｅｌｌｉｆｅｒａ）毒腺中提取总ＲＮＡ,通过ＲＴ－ＰＣＲ方法扩增得到了蜂毒肽前体蛋白的ｃＤＮＡ。扩增产物克隆到ｐＴ７Ｂｌｕ－Ｔ载体,再进一步将插入片段酶切连接到ＰＵＣ１１８载体上,构建了与β－半乳糖苷酶部分序列相融合的蜂毒肽前体蛋白表达体并转化大肠杆菌ＪＭ１０９。     

Novel cytotoxic peptides from the tropical marine cyanobacteriumHormothamnion enteromorphoides 1. Discovery,isolation and initial chemical and biological characterization of the hormothamnins from wild and cultured material

A Caribbean cyanobacterium,Hormothamnion enteromorphoides, was found to produce a complex mixture of ichthyotoxic peptides, perhaps explaining the apparent absence of predation upon these potentially palatable life forms. Bioassay-guided fractionation was used to isolate these toxic and antimicrobial natural products, and a variety of techniques including HR FAB mass spectrometry, 2D-NMR, traditional hydrolysis-amino acid analysis, and several chemical reactions were used to define the basic structural features of the major peptide, hormothamnin A. Hormothamnin A is a cyclic undecapeptide containing six common and five uncommon or new amino acid residues. HPLC analyses indicate that the relative proportions of these peptide natural products remain relatively constant between different collection locations and years, however, they do vary seasonally. Clonal isolates of this cyanobacterium in culture produce the full spectrum of toxic peptides.     

Aspects of measurement and analysis of regulatory peptides

Summary Although almost all methods of mass measurement of regulatory peptides still depend on the high affinity antibody, the traditional Yalow and Berson radioimmunoassay technique is becoming outdated. Pure monoclonal antibodies allow excess antibody two site assay techniques with a variety of different labels (preferentially non-radioactive) of great sensitivity and speed. The large amounts of particular monoclonal antibodies available allow several different laboratories to use the same reagents and have increased comparability. Unfortunately many regulatory peptides exist in multiple molecular forms and attention must be paid to antibody region specificity. Improved methods of extraction of regulatory peptides from plasma tissue allow more accurate quantitation. New techniques for rapid high resolution chromatography make distinction of different molecular forms much easier than hitherto. Better education in techniques and/or attention to inter-assay standards are necessary to improve the comparability of regulatory peptide measurement in the future.     

Precursors to regulatory peptides: their proteolytic processing

Summary Precursors to regulatory peptides undergo maturation processes which include protelytic processing. The enzymes involved in this process remove the hydrophobic peptide located at the amino-terminus of the precursor. Endoprotease cleavage also occurs at single and two adjacent basic residues, this is followed by a removal of basic residues located at the C-terminus of the peptides by a carboxypeptidase-like enzyme.     

Cell-penetrating peptides: tools for intracellular delivery of therapeutics

The main problem of therapeutic efficiency lies in the crossing of cellular membranes. Therefore, significant effort is being made to develop agents which can cross these barriers and deliver therapeutic agents into cellular compartments. In recent years, a large amount of data on the use of peptides as delivery agents has accumulated. Several groups have published the first positive results using peptides for the delivery of therapeutic agents in relevant animal models. These peptides, called cell-penetrating peptides (CPPs), are short peptides (fewer than 30 residues) with a net positive charge and acting in a receptor- and energy-independent manner. Here, we give an extensive review of peptide-mediated delivery systems and discuss their applications, with particular focus on the mechanisms leading to cellular internalization.Received 14 March 2005; received after revision 25 April 2005; accepted 28 April 2005     

Mass spectrometry-based proteomics in the life sciences

Over the last 20 years, mass spectrometrybased proteomics has become an indispensable tool in the cellular and molecular life sciences. This has been enabled by the soft ionisation techniques of electrospray and matrix-assisted laser desorption-ionisation, which allow the gentle ionisation and vaporisation of large, thermally labile biomolecules. Innovative instrumentation designs and biochemical strategies have brought success in the large-scale identification and quantification of proteins, as well as the characterisation of their complexes and post-translational modifications. This review describes the instrumentation used for proteomics research. It presents an overview of the current applications of mass spectrometry-based proteomics to the cellular and molecular life sciences, and discusses challenges that exist for research in the future.Received 7 January 2005; accepted 27 January 2005     

多肽生化和手性药物的研究

综述了我研究室近年来在多肽生化和手性药物领域中的研究进展：①进行了抗癌药物 Ａ Ｍ Ｄ 类似物的全新设计、化学全合成、与 Ｄ Ｎ Ａ 结合活性和构效关系研究； ②研究了新皮啡肽类似物及其自旋标记衍生物和孤儿受体的天然配基孤啡肽的构效关系；③进行了 Ｄ Ｎ Ａ 结合蛋白结构域：锌指及其突变体的合成及与寡聚核苷酸结合能力的研究； ④将稳定的氮氧自由基标记肽类药物血管紧张素Ⅱ，建立了一种使生物活性肽带上自旋标记的简便方法， 用以研究肽类药物与受体间的相互作用及自由基对肽类药物生理活性的影响；⑤为解决许多手性药物合成中的难题，开发了一个构筑手性季碳的新方法； ⑥研究了手性催化剂作用下二乙基锌对醛的不对称加成反应，为手性药物的研制提供了新反应     

表皮葡萄球菌RP62A和ATCC12228差异蛋白的进化速率分析

表皮葡萄球菌作为一种主要的医院感染病原体越来越受到关注.其致病的主要因素在于致病株进化出了能在医疗器械表面生成生物膜的机制.为了研究表皮葡萄球菌的致病性,本文比较了致病株和非致病株之间蛋白质组表达的差异性,揭示了与生物膜形成相关的基因表现出差异表达的趋势.结合同源蛋白对进化速率的分析结果显示,表皮葡萄球菌的蛋白表达丰度和进化速率呈负相关的趋势.同源蛋白对进化速率的分析结果还显示,表皮葡萄球菌蛋白质组中一部分蛋白受到了较强的负选择压力,是微生物体中保守的部分;还有一部分受到的选择压力较小,对生物体应对外界环境做出遗传改变从而产生相应的表型有很大帮助.     

酪丝缬肽的结构修饰及其稳定性和安全性的初步评价

通过固相多肽法成功合成了酪丝缬肽(YSV)以及其结构修饰物(Z-GP-YSV-NH2),总收率分别为45.3%和52.3%。分别研究了YSV和Z-GP-YSV-NH2在PBS缓冲液、DMEM培养基和大鼠血浆中的稳定性,发现两者在PBS和DMEM中都很稳定,但YSV在大鼠血浆中温浴2 h后基本降解完全;而Z-GP-YSV-NH2温浴12h后只降解67%。还评价了YSV和Z-GP-YSV-NH2对人胚肾细胞293的细胞毒性,发现两者的细胞毒性都很低。当剂量为200μg/mL时,其增殖抑制率分别为20%和10.3%。