定向置换细菌和真菌的结构域,推理设计肽类抗生素

由微生物产生的次级代谢产物肽,无论在化学结构或生物活性上都具有多样性.这类代谢产物包括着抗生素、酶抑制剂、植物或动物毒素和免疫抑制剂等,在医学、农学和生物学研究上都有重要的意义.早己鉴定过两种氨基酸拼入活性多肽的机理.譬如,多环抗生素是由基因编码的肽前体核糖体合成的,而这些肽前体经复合的翻译后加工(Post-translational processing)被广泛加以修饰.另一些肽则是以一种非核糖体的机理,在蛋白质的模板上合成的.这种合成提供了结构多种多样的肽次级代谢产物,它们可由线状的、环状的或分支的肽链组成;可含有D型-、羟基-或N甲基-氨基酸,这些氨基酸又可做酰基化或糖基化等修饰.     

先导化合物结构优化策略(七)——肽类分子结构修饰与改造

多肽分子大多为天然的内源性配体,与受体的亲和力强,选择性好,是一类比较容易成为先导化合物及药物的分子。许多药物都是从多肽分子进行结构优化改造而得,如降压药物卡托普利、抗丙肝药物特拉匹韦等。目前多肽分子开发面临的主要问题包括稳定性差、半衰期短、血浆清除率高等低成药性缺陷;通常只能注射使用,患者的依从性较差;生产工艺复杂,生产成本较高。因此,对多肽分子进行合理的修饰和改造既可以降低肽类分子的生产成本,又可以改善肽类分子的成药性。本文从改善肽类分子成药性的角度综述了肽类分子结构修饰与改造策略,根据是否对肽链骨架进行修饰,将修饰策略分为两类:一类是针对肽链骨架的改造,包括非天然氨基酸修饰、伪肽化策略、逆肽策略、环化策略、末端结构修饰等;另一类是在多肽骨架不变的基础上,引入其他基团进行结构优化和性能改造,包括高级脂肪酸修饰、聚乙二醇修饰、蛋白融合策略、胆固醇修饰等。     

与P物质有关的六肽的构象-活性关系

十一肽P物质(Arg-Pro-Lys-Pro-Gin-Gln-Phe-Phe-Gly-Leu-Met-NH_2)(SP)属速激肽类物质。这些多肽有两种受体:SP-P 存在于豚鼠回肠壁(GPI),SP-E存在于大鼠的输精管(RVD)中。本文先用Glu 代替Gln~6以增强N-端的亲水性,再引入一些官能团以增加对SP 受体亚类的亲和性,然后用Pro 取代Gly~9。这种修饰目的在于确定这种理论上有利于折叠式构象的变化,是否影响SP 受体的生物活性和选     

肽类药物国内外研究进展

多肽是指由氨基酸组成，氨基酸用肽键连接而成的一类化合物，其在连接方式上与蛋白质相同，通常将少于100个氨基酸组成的称为多肽，而由100个以上氨基酸组成的为蛋白质。但在某些性质方面多肽又有别于蛋白质，如其空间结构较简单、稳定性较高、免疫原性较低或无免疫原性等。所有生物细胞都能合成多肽，同时也受各种多肽的调节。多肽的生物活性广泛而重要，     

合成多肽药物质量控制研究探讨

合成多肽药物作为一种介于小分子和生物制品之间的特殊药物形式，既有小分子药物稳定性好和纯度高的特点，又具有生物药物生物活性高和毒副作用低的优势，越来越受到制药行业的关注。多肽是由多个氨基酸通过酰胺键共价连接而成，由于氨基酸异构和侧链修饰导致多肽药物与生物制品一样存在异质性的特点，而化学合成工艺复杂多样，杂质谱的鉴定相比小分子药物更为困难。因此合成多肽药物的质量控制既要满足化学合成药物的质量控制要求，又要兼顾生物制品的特点，对于制药行业存在很大挑战。文章结合国内外合成多肽药物药学研究的相关指导原则及研究进展，从多肽的鉴别、含量测定、杂质谱分析等方面出发，对合成多肽药物的质量控制进行全面的探讨和分析，以期为后续合成多肽类药物的质量控制研究提供参考。     

人ⅡA型磷脂酶A_2衍生肽P26抗菌作用与结构修饰的研究

目的比较人ⅡA型磷脂酶A2(phospholipase A2,PLA2)C末端衍生的多肽和经修饰后的环肽对不同细菌在体外的杀菌效应,了解其作用机制。方法根据人ⅡA型PLA2氨基酸顺序C末端26个氨基酸残基,合成直链肽P26,同时将其修饰为环肽P26。采用琼脂铺板计数法,将不同浓度的两种多肽分别与6种细菌在37℃孵育2 h,然后铺板并置于37℃恒温箱培养18～24 h,记录每一琼脂板上的菌落数(CFU),并计算多肽作用后的杀菌率。结果两种多肽对革兰阳性菌金黄色葡萄球菌、枯草杆菌和炭疽杆菌有较强的杀菌活性,环肽P26的作用比直链肽P26强4倍左右;对革兰阴性菌大肠杆菌、变形杆菌和绿脓杆菌的杀菌作用较弱,而环肽P26的作用更弱。结论衍生自人ⅡA型PLA2 C末端26个氨基酸残基的肽P26和修饰得到的环肽P26对G+菌有较强的杀菌活性,经修饰成环肽后作用更强,可能与环肽分子带的正电荷更集中有关。     

含APD环缩酚酸肽类天然产物的研究进展

含有4-氨基-2,4-戊二烯酸的环缩酚酸肽类天然产物是一类包含酯键的环状多肽次级代谢产物,通常具有抗肿瘤、抗病原体等生物活性。此文根据国内外最新研究现状,介绍了这类物质的来源、结构、制备现状、生物活性以及开发前景。     

利用高分辨质谱结合生物信息学分析预测注射用心肌肽中的生物活性肽

目的:基于高分辨率质谱多肽鉴定技术，分析注射用心肌肽中多肽类物质组成，并采用生物信息技术预测多肽的生物活性。方法:注射用心肌肽净化除盐后，以Orbitrap Fusion Lumos三合一高分辨质谱系统对混合多肽进行鉴定，利用BIOPEP-UWM数据库工具预测肽兰克评分≥0.80的肽段的潜在生物活性。结果:注射用心肌肽经分析鉴定出799条肽段，分别来源于70种蛋白质，主要为肌动蛋白、血红蛋白亚基α、ADP/ATP移位酶3等。肽兰克评分≥0.80的10条肽段中具有丰富的血管紧张素转化酶抑制活性(ACEi)、二肽基肽酶-4抑制活性(DPP4i)及抗氧化活性(AO)氨基酸片段。结论:超高效液相色谱-高分辨质谱联合技术及生物信息技术为多组分生化药分析鉴定及活性预测提供便捷手段。注射用心肌肽的心肌保护等作用的物质基础可能源于其中的多肽影响血管紧张素转化酶、抗氧化及二肽基肽酶-4等多种生物活性。     

新型三维氨基酸结构描述符的研究及其在多肽QSAR中的应用

目的建立一套新型的氨基酸残基结构描述符，用此描述符进行多肽的定量构效关系研究。方法通过理论计算得出167个氨基酸结构描述参数，再对167个参数进行主成分分析，从中提取出与氨基酸残基的三维结构性质密切相关的三个结构描述参数c₁，c₂，c₃-scales。然后采用G/PLS算法，建立氨基酸残基结构描述符c-scales与多肽生物活性之间的函数关系，确定多肽的QSAR模型。结果用此方法建立的苦味二肽、ACE抑制剂二肽和缓激肽增效剂五肽的QSAR模型，均有较好的拟合和预测能力，交叉验证相关系数和相关系数都在0.70以上。结论氨基酸残基结构描述符c-scales能定量的描述氨基酸的三维结构性质，而且利用此描述符能建立预测能力较强的多肽QSAR模型。     

多肽激素类似物的设计

生物活性多肽的领域近年来有很大扩展,尤其是发现了很多具有激素功能的多肽,也可以作为中枢及周围神经系统的神经递质或者神经调节剂。现已弄清,多肽是使身体各部分形成整体和精确调节机体各种功能的主要物质。生物活性多肽的氨基酸顺序一经确定,不仅为化学家打开了合成天然分子的道路,而且可以合成其他类似物。最近,多肽合成技术的进步已使小分子和中等分子多肽的合成成为常规工作,而且有可能合成大分子肽甚至小分子蛋白质。合成生物活性多肽类似物有几个理由。首先,我们要了解多肽-受体相互作用的本质,因为多肽具有很多能够与受体发生相互     

Efficient macrocyclization for cyclicpeptide using solid-phase reaction

Cyclicpeptides are important targets in peptide synthesis because of their interesting biological properties. Constraining highly flexible linear peptides by cyclization is one of the mostly widely used approaches to define the bioactive conformation of peptides. Cyclic peptides often have increased receptor affinity and metabolic stability over their linear counterparts. We carried out virtual screening experiment via docking in order to understand the interaction between HLE-Human Leukocyte Elastase and ligand peptide and to identify the sequence that can be a target in various ligand peptides. We made cyclic peptides as a target base on Met-Ile-Phe sequence having affinity for ligand and receptor active site docking. There are three ways to cyclize certain sequences of amino acids such as Met-Ile-Phe-Gly-Ile. First is head-to-tail cyclization method, linking between N-terminal and C-terminal. Second method utilizes amino acid side chain such as thiol functional group in Cys, making a thioether bond. The last one includes an application of resin-substituted amino acids in solid phase reaction. Among the three methods, solid phase reaction showed the greatest yield. Macrocyclization of Fmoc-Met-Ile-Phe-Gly-Ile-OBn after cleavage of Fmoc protection in solution phase was carried out to give macrocyclic compound 5 in about 7% yield. In the contrast with solution phase reaction, solid phase reaction for macrocyclization of Met-Ile-Phe-Gly-Ile-Asp-Tentagel in normal concentrated condition gave macrocyclic compound 7 in more than 35% yield.     

Unusual amino acids: synthesis and introduction into naturally occurring peptides and biologically active analogues

This review covers our recent advances in the synthesis of unusual amino acids in optically pure form, and their introduction into naturally occurring peptides with specific biological properties, or into modified bioactive peptides, aiming to obtain analogues displaying enhanced performances in term of activity, bioavailability and resistance to enzymatic hydrolysis.     

Development of conformationally restricted analogues of bradykinin and somatostatin using constrained amino acids and different types of cyclization

The structure-based design of peptide drugs requires the knowledge of the bioactive conformation. Studies on this receptor-bound 3D structure require linear or cyclic analogues with strongly reduced flexibility, but high biological activity, since only analogues with retained potency have preserved the bioactive conformation. Constrained amino acids containing double bonds or bulky substituents at the N(alpha)-, C(alpha)- and C(beta)-atom as well as at the aromatic ring atom were successfully applied to obtain potent and stable analogues of bradykinin and somatostatin, which due to their restricted conformation were suitable objects for conformational studies. Besides the generation of constrained cyclic analogues with improved biological and pharmacological properties, cyclic peptides were used as convenient models for the study of turn formations. Cyclization of the linear peptide bradykinin was performed by linking the N-terminus and the C-terminus, and in both bradykinin and somatostatin by cyclization using the amino acid side chains and by backbone cyclization. The later requires the introduction of N(alpha)-functionalised amino acids for ring closure which can be performed either through incorporation of N(alpha)-functionalised amino acids or dipeptide building units. Conformational analysis of a cyclic bradykinin analogue by means of NMR-studies together with molecular dynamics simulation led to a quasicyclic 3D structure with two turns and together with other 3D structures provided a pharmacophore model of bradykinin antagonists.     

N- and Calpha-methylation in biologically active peptides: synthesis, structural and functional aspects

Numerous backbone constraints can be used to develop pseudopeptides or pseudomimetics of biologically active peptides. Among those, N- and Calpha-methyl amino acids that can be incorporated by solid-phase peptide synthesis in a bioactive sequence represent important tools to restrict phi and psi angles of peptide backbone. This review will focus on the chemical syntheses of N- and Calpha-methyl amino acids, their effects on peptide conformation and structure, and their role on the peptide stability towards enzymatic degradation and on the biological activities of the resulting analogues.     

Bioactive proteins and peptides from food sources. Applications of bioprocesses used in isolation and recovery

There are many examples of biologically active food proteins, with physiological significance beyond the pure nutritional requirements that concern available nitrogen for normal growth and maintenance. Moreover, there are many physiologically active peptides, derived by protease activity from various food protein sources; however, relationships between structural properties and functional activities have not been completely elucidated. Many bioactive peptides have in common structural properties that include a relatively short peptide residue length (e.g. 2-9 amino acids), possessing hydrophobic amino acid residues in addition to proline, lysine or arginine groups. Bioactive peptides are also resistant to the action of digestion peptidases. Antihypertensive peptides, known as Angiotensin I converting enzyme (ACE) inhibitors have been derived from milk, corn and fish protein sources. Peptides with opioid activities are derived from wheat gluten or casein, following digestion with pepsin. Exorphins, or opioid peptides derived from food proteins such as wheat and milk (e.g. exogenous sources) have similar structure to endogenous opioid peptides, with a tyrosine residue located at the amino terminal or bioactive site. Immunomodulatory peptides derived from tryptic hydrolysates of rice and soybean proteins act to stimulate superoxide anions (reactive oxygen species-ROS), which triggers non-specific immune defense systems. Antioxidant properties that prevent peroxidation of essential fatty acids have also been shown for peptides derived from milk proteins. The addition of a Leu or Pro residue to the N-terminus of a His-His, dipeptide will enhance antioxidant activity and facilitate further synergy with non-peptide antioxidants (e.g. BHT). We also show herein, that the tryptic digests of casein yielding caseinophosphopeptides exhibits both hydrophilic and lipophilic antioxidant activity due to both metal ion sequestering and quenching of ROS. The separation and purification of bioactive peptides which will involve development of automated and continuous systems is an important field for Food chemists. Much effort has been given to develop selective column chromatography methods that can replace batch methods of salting out, or using solvent extraction to isolate and purify bioactive peptides. Advances here will enable recovery of bioactive peptides with minimal destruction thus enabling utilization by returning these active peptides to functional food or specific nutraceutical applications.     

Technological options for the production of health-promoting proteins and peptides derived from milk and colostrum

Milk proteins are known to exert a wide range of nutritional, functional and biological activities. Apart from being a balanced source of valuable amino acids, milk proteins contribute to the consistency and sensory properties of various dairy products. Furthermore, many milk proteins possess specific biological properties which make them potential ingredients of health-promoting foods. These properties are attributed to both native protein molecules and to physiologically active peptides encrypted in the protein molecules. Considerable progress has been made over the last twenty years in technologies aimed at separation, fractionation and isolation in a purified form of many interesting proteins occurring in bovine colostrum and milk. Industrial-scale methods have been developed for native whey proteins such as immunoglobulins, lactoferrin, lactoperoxidase, alpha-lactalbumin and beta-lactoglobulin. Their large-scale manufacture and commercial exploitation is still limited although validated research data about their physiological health benefits is rapidly accumulating. Promising product concepts and novel fields of use have emerged recently, and some of these molecules have already found commercial applications. The same applies to bioactive peptides derived from different milk proteins. Active peptides can be liberated during gastrointestinal digestion or milk fermentation with proteolytic enzymes. Such peptides may exert a number of physiological effects in vivo on the gastrointestinal, cardiovascular, endocrine, immune, nervous and other body systems. However, at present the industrial-scale production of such peptides is limited by a lack of suitable technologies. On the other hand, a number of bioactive peptides have been identified in fermented dairy products, and there are already a few commercial dairy products enriched with blood pressure-reducing milk protein peptides. There is a need to develop methods to optimise the activity of bioactive peptides in food systems and to enable their optimum utilisation in the body. This review highlights existing modern technologies applicable for the isolation of bioactive native proteins and peptides derived from bovine colostrum, milk and cheese whey, and discusses aspects of their current and potential applications for human nutrition and promotion of human health.     

Food-derived bioactive peptides--opportunities for designing future foods

Dietary proteins are known to carry a wide range of nutritional, functional and biological properties. Nutritionally, the proteins are a source of energy and amino acids, which are essential for growth and maintenance. Functionally, the proteins contribute to the physicochemical and sensory properties of various protein-rich foods. Furthermore, many dietary proteins possess specific biological properties which make these components potential ingredients of functional or health-promoting foods. Many of these properties are attributed to physiologically active peptides encrypted in protein molecules. Particularly rich sources of such peptides are milk and egg, but they are also found in meat of various kinds as well as many plants. These peptides are inactive within the sequence of parent protein and can be released during gastrointestinal digestion or food processing. Depending on the amino acid sequence, these peptides may exert a number of different activities in vivo, affecting, e.g., the cardiovascular, endocrine, immune and nervous systems in addition to nutrient utilization. There is increasing commercial interest in the production of bioactive peptides from various sources. Industrial-scale production of such peptides is, however, hampered by the lack of suitable technologies. Bioactive peptides can also be produced from milk proteins through fermentation of milk, by starters employed in the manufacture of fermented milks or cheese. In particular, antihypertensive peptides have been identified in fermented milk, whey and ripened cheese. A few of these peptides have been commercialised in the form of fermented milks. There is a need to develop technologies which retain or even enhance the activity of bioactive peptides in food systems. Also, it is essential to study the optimum utilization of such peptides during passage through the gastrointestinal tract.     

Design and synthesis of non-proteinogenic amino acids and secondary structures of their peptides

Replacement of the alpha-hydrogen atom of L-alpha-amino acids with an alkyl substituent results in alpha,alpha-disubstituted amino acids. The modification changes the properties of the amino acids. Incorporation of alpha,alpha-disubstituted amino acids into oligopeptides restricts the conformational freedom of their peptides. The author developed a synthetic route for optically active alpha-ethylated alpha,alpha-disubstituted amino acids using chiral cyclic 1,2-diol, and disclosed that the preferred conformation of peptides composed of chiral alpha-ethylated disubstituted amino acids is a fully planar conformation, whereas that of chiral alpha-methylated disubstituted amino acids is a 3(10)-helical structure. Furthermore, the author designed and synthesized two chiral cyclic alpha,alpha-disubstituted amino acids, i.e., (3S,4S)-1-amino-3,4-di(methoxy)cyclopentanecarboxylic acid {(S,S)-Ac5cdOM}, and (1R,6R)-8-aminobicyclo-[4.3.0]non-3-ene-8-carboxylic acid {(R,R)-Ab5,6=c}. They do not have a chiral center at the alpha-position, but do have chiral centers on the side-chain cyclopentane or the bicyclic skeleton. The preferred secondary structure of the (S,S)-Ac5cdOM homopeptides was the left-handed (M) 3(10)-helical structure (hexapeptide) and the left-handed (M) alpha-helical structure (octa- and decapeptides), while that of the (R,R)-Ab5,6=c hexapeptide was both the right-handed (P) and left-handed (M) 3(10)-helices. These results indicate that the side-chain chiral centers affect the secondary structure of their peptides, and the side-chain chiral environment is important for the control of the helical-screw direction of peptides.     

Sequence-dependence of secondary structure formation II*

The conformational influence of the insertion of two guest amino acids into a homooligopeptide consisting of L-Val residues was investigated by circular dichroism and infrared spectroscopy. L-Ala, L-Ile and L-Leu were chosen as guest amino acids and their β-structure disrupting properties studied with respect to size and symmetry of their side-chains. These studies were complemented by investigations concerning the aggregation behaviour of the various peptides by means of gel permeation chromatography. It is shown that the tendency of the various peptides to form β-structures and to aggregate increases with increasing similarity in the spatial size of the side-chains between guest and host amino acids. The impact of those results on the prediction of peptide secondary structure as well as their importance for peptide synthesis is briefly discussed.