合成生物学指导下芽孢杆菌合成环脂肽的研究进展*

近年来,合成生物学借助工程化在人工生命系统的设计与构建方面取得了长足进展,特别是“细胞工厂”的开发和应用为天然产物的合成带来了深刻变革。环脂肽是一类新型的天然表面活性剂,因其特殊的结构和功能亦可作为抗生素使用。目前,合成环脂肽最理想的微生物底盘是芽孢杆菌。因此,许多研究者致力于通过合成生物学技术来提升芽孢杆菌作为环脂肽细胞工厂的性能。首先,对芽孢杆菌中环脂肽的非核糖体肽合成途径进行概述;其次,重点介绍与环脂肽合成相关的调控因子;再次,从底盘细胞的选择、基因编辑工具的开发、合成路径的优化及发酵过程的优化等四个方面对合成生物学指导下环脂肽的相关研究进展进行总结;最后,讨论环脂肽合成中可能存在的挑战,并就未来研究趋势进行展望,以期为高效环脂肽细胞工厂的开发提供参考。     

放线菌来源的羊毛硫肽研究进展

羊毛硫肽(lanthipeptide)是一类由核糖体合成并经翻译后修饰的含羊毛硫氨酸或β-甲基羊毛硫氨酸的多肽。近年来,放线菌来源的羊毛硫肽因其突出的抗菌活性和罕见的生物活性而备受关注。本文重点对放线菌来源的不同类型的羊毛硫肽的结构特征及其特性进行了综述,讨论了生物或化学方法修饰天然羊毛硫肽和基因组挖掘发现结构新颖的羊毛硫肽在开发符合实际应用需求的放线菌来源的羊毛硫肽中的应用,并对放线菌来源的羊毛硫肽的应用潜力进行了总结和展望。     

细胞穿透肽应用的研究进展

细胞穿透肽是一类具有特殊细胞膜穿透作用的短肽,研究发现其为药物大分子物质的细胞内转运提供了有力工具,现已得到了广泛关注和大量研究.一方面,近年来发现,细胞穿透肽的应用面临着药物释放率、代谢降解、细胞系的分化状态和Rho-GTPases活性的依赖性等问题,从而阻碍了其研究开发的进程.另一方面,细胞穿透肽在肿瘤治疗、抗菌和抗炎靶向药物等方面的研究却取得了显著的进展,甚至进入了临床前和临床研究阶段.本文围绕细胞穿透肽在应用开发中所面临的挑战和机遇两方面,综述了近期的研究进展,并对细胞穿透肽在新领域中的应用进行了展望.     

生物活性肽

生物活性肽是对生物机体的生命活动有益或具有生理作用的肽类化合物,简述了生物活性肽的分类、特点、作用机理以及开发应用。     

肽抗生素hPAB-β3拷贝工程菌的发酵与重组蛋白纯化

肽抗生素 (peptideantibiotics)是近年来发现的生物体基因编码的具有抗微生物活性的小肽 ,在临床病原菌对常用抗生素耐药性日益严重的今天 ,人们希望将肽抗生素开发为一种新型的抗感染制剂。为大量制备人源肽抗生素hPAB β ,本研究在前期对筛选出的 3拷贝优势工程菌的摇瓶发酵条件进行摸索的基础上 ,采用 3.7L发酵罐对发酵pH值、溶氧条件 (PO2 )和补料时机、方式等参数进行优化。同时 ,对表达产物进行了如下纯化 :利用Ni NTA亲和层析捕获 3拷贝融合蛋白 ,继而用 2mol/L羟胺裂解 ,产物用SOURCE 30RPC填料进行反相层析 ,目标肽经GSH…     

海洋细菌活性蛋白、活性肽研究的若干新进展

海洋蕴藏着丰富的生物活性资源,近年来海洋细菌活性蛋白及活性肽的研究在资源开发与利用领域中很受关注。本文首先分析了海洋细菌来源的各种活性蛋白及活性肽在不同层面上的生物学活性特征,其次对海洋活性蛋白及活性肽的获得方式及其在医药学、工业、水产养殖业及环境保护等领域中的应用现状与潜力作简要介绍,最后对该领域研究前景进行了展望。     

乳清蛋白及其生物活性肽血压调节功能研究进展

乳清蛋白既是优质蛋白来源，也是抗高血压生物活性肽的理想来源，开发具有调节血压功效的蛋白多肽类产品对未来临床高血压防治将发挥重要作用。文章综述了乳清蛋白生物活性肽血压调节功能的基本机制和研究现状，并对其未来发展趋势和应用前景进行了讨论。     

Merck公司开始hANP的临床试验

美国Merck公司开始进行人心房性钠利尿肽的临床试验。 ANP是由心脏中分泌的、在肾功能和血压等体液调节中起重要作用的肽激素。世界上现有近30家公司从事高血压药物的开发,其中也包括ANP的研究开发。最近据报道,ANP对心脏功能不全也有效,所以人们对它寄予厚望。     

噬菌体展示技术筛选bFGF模拟短肽

目的：通过噬菌体展示技术筛选得到与FGFR结合的bFGF模拟短肽,为bFGF肽类抑制剂的研发提供实验基础。方法：以Balb/c 3T3细胞为靶标,以COS-7细胞作消减,对噬菌体随机七肽库进行4轮生物淘洗,再采用ELISA检测单克隆噬菌体对Balb/c 3T3亲和性和特异性,选取阳性克隆进行DNA测序分析。结果：从富集的噬菌体中获得12个阳性克隆,获得一组疏水性七肽及共同基序PR。结论：利用肽类新药开发的重要工具--噬菌体展示技术,得到2段bFGF的受体结合模拟肽,可望作为bFGF抑制剂的先导肽。     

花生肽功能特性研究进展

花生肽因其丰富的营养价值、卓越的功能特性、独特的吸收机制,近来备受关注。综述了国内外对花生肽的功能特性的研究进展,分析问题并提出展望,为花生肽的合理开发提供指导。     

Characterization and production of multifunctional cationic peptides derived from rice proteins

Food proteins have been identified as a source of bioactive peptides. These peptides are inactive within the sequence of the parent protein and must be released during gastrointestinal digestion, fermentation, or food processing. Of bioactive peptides, multifunctional cationic peptides are more useful than other peptides that have specific activity in promotion of health and/or the treatment of diseases. We have identified and characterized cationic peptides from rice enzymes and proteins that possess multiple functions, including antimicrobial, endotoxin-neutralizing, arginine gingipain-inhibitory, and/or angiogenic activities. In particular, we have elucidated the contribution of cationic amino acids (arginine and lysine) in the peptides to their bioactivities. Further, we have discussed the critical parameters, particularly proteinase preparations and fractionation or purification, in the enzymatic hydrolysis process for producing bioactive peptides from food proteins. Using an ampholyte-free isoelectric focusing (autofocusing) technique as a tool for fractionation, we successfully prepared fractions containing cationic peptides with multiple functions.     

In silico analysis of the large and small subunits of cereal RuBisCO as precursors of cryptic bioactive peptides

Ribulose-1,5-bisphosphate carboxylase (RuBisCO) is a plant metabolic enzyme and the most abundant protein on earth, but remains largely underutilized in the food system. Bioinformatics analysis was conducted to evaluate the prospects of RuBisCO from widely cultivated cereals (rice, barley, wheat, oat, sorghum, corn) as sources of bioactive peptides, and results were compared to commonly consumed proteins. The large and small RuBisCO subunits were found to contain several bioactive peptides with biological functions relevant to the management and treatment of diabetes, cardiovascular and neurodegenerative diseases, specifically inhibition of angiotensin converting enzyme and dipeptidyl peptidase-IV, antioxidative property and activation of ubiquitin-mediated proteolysis. Due to high sequence homology, there was negligible difference in occurrence frequency of bioactive peptides within large RuBisCO subunits unlike small subunits, which produced more diverse profiles of bioactive peptides among the cereals. The cereal RuBisCO displayed similar or better prospects as other food proteins except milk proteins, thereby providing cheaper and sustainable precursors of bioactive peptides. Simulated enzymatic hydrolysis of RuBisCO subunits indicated that thermolysin and papain had preferred cleavage patterns for releasing the cryptic peptides compared to gastrointestinal proteases. These findings will contribute towards utilization of RuBisCO as alternative sources of peptide-based nutraceuticals for human health promotion.     

Bioactive peptides derived from human milk proteins — mechanisms of action

Human milk contains a multitude of bioactive proteins with very diverse functions, which are beneficial for the rapidly growing neonate. The large variety of bioactivities is accomplished by the combination of bioactive proteins per se and gastrointestinal release of bioactive peptides derived from them. The bioactivities exerted by these peptides include enhancement of mineral absorption, immunomodulation, opioid, antihypertensive and antimicrobial activities. Notably, several of the activities are not attributed to the parental proteins, but exclusively to released bioactive peptides. This article reviews studies on bioactive peptides derived from major human milk proteins, such as caseins, α-lactalbumin and lactoferrin, during gastrointestinal digestion. Studies of bovine milk counterparts are also cited as a comparison.     

Marine natural product peptides with therapeutic potential: Chemistry,biosynthesis, and pharmacology

The oceans are a uniquely rich source of bioactive metabolites, of which sponges have been shown to be among the most prolific producers of diverse bioactive secondary metabolites with valuable therapeutic potential. Much attention has been focused on marine bioactive peptides due to their novel chemistry and diverse biological properties. As summarized in this review, marine peptides are known to exhibit various biological activities such as antiviral, anti-proliferative, antioxidant, anti-coagulant, anti-hypertensive, anti-cancer, antidiabetic, antiobesity, and calcium-binding activities. This review focuses on the chemistry and biology of peptides isolated from sponges, bacteria, cyanobacteria, fungi, ascidians, and other marine sources. The role of marine invertebrate microbiomes in natural products biosynthesis is discussed in this review along with the biosynthesis of modified peptides from different marine sources. The status of peptides in various phases of clinical trials is presented, as well as the development of modified peptides including optimization of PK and bioavailability.     

Physico-chemical,Sensory and Toxicity Characteristics of Dipeptidyl Peptidase-IV Inhibitory Peptides from Rice Bran-derived Globulin Using Computational Approaches

Rice processing industry released an enormous amount of the rice bran which is underutilized. Rice bran contains various proteins that can be used for the production of bioactive peptides. These bioactive peptides might be suitable ingredients for the development of functional food products. The objective of this study was to explore the potential of rice bran-derived globulin proteins as a suitable precursor of bioactive peptides with especially reference to dipeptidyl peptidase IV (DPP-IV) inhibitory peptides. The various computational approaches (BLAST, BIOPEP, PeptideRanker, PepDraw, Pepcalc, and ToxinPred) were used to predict the potential of the globulin proteins. Ficain protease majorly released the DPP-IV inhibitory peptides from rice bran-derived globulin proteins as compared with other proteases used in this study. Furthermore, primary structure, physico-chemical, sensory, and allergic characteristics of the theoretically release bioactive DPP-IV inhibitory peptides were also studied. The result of this study provides a theoretical basis for the development of rice bran globulin proteins as a suitable source for the generation of bio-functional ingredients for glycaemic management and further demonstrates the usefulness of computational approaches.     

Towards the Improved Discovery and Design of Functional Peptides: Common Features of Diverse Classes Permit Generalized Prediction of Bioactivity

The conventional wisdom is that certain classes of bioactive peptides have specific structural features that endow their particular functions. Accordingly, predictions of bioactivity have focused on particular subgroups, such as antimicrobial peptides. We hypothesized that bioactive peptides may share more general features, and assessed this by contrasting the predictive power of existing antimicrobial predictors as well as a novel general predictor, PeptideRanker, across different classes of peptides.We observed that existing antimicrobial predictors had reasonable predictive power to identify peptides of certain other classes i.e. toxin and venom peptides. We trained two general predictors of peptide bioactivity, one focused on short peptides (4–20 amino acids) and one focused on long peptides ( amino acids). These general predictors had performance that was typically as good as, or better than, that of specific predictors. We noted some striking differences in the features of short peptide and long peptide predictions, in particular, high scoring short peptides favour phenylalanine. This is consistent with the hypothesis that short and long peptides have different functional constraints, perhaps reflecting the difficulty for typical short peptides in supporting independent tertiary structure.We conclude that there are general shared features of bioactive peptides across different functional classes, indicating that computational prediction may accelerate the discovery of novel bioactive peptides and aid in the improved design of existing peptides, across many functional classes. An implementation of the predictive method, PeptideRanker, may be used to identify among a set of peptides those that may be more likely to be bioactive.     

Biological activities and potential health benefits of bioactive peptides derived from marine organisms

Marine organisms have been recognized as rich sources of bioactive compounds with valuable nutraceutical and pharmaceutical potentials. Recently, marine bioactive peptides have gained much attention because of their numerous health beneficial effects. Notably, these peptides exhibit various biological activities such as antioxidant, anti-hypertensive, anti-human immunodeficiency virus, anti-proliferative, anticoagulant, calcium-binding, anti-obesity and anti-diabetic activities. This review mainly presents biological activities of peptides from marine organisms and emphasizing their potential applications in foods as well as pharmaceutical areas.     

Peptidomic identification and biological validation of neuroendocrine regulatory peptide-1 and -2

Recent advances in peptidomics have enabled the identification of previously uncharacterized peptides. However, sequence information alone does not allow us to identify candidates for bioactive peptides. To increase an opportunity to discover bioactive peptides, we have focused on C-terminal amidation, a post-translational modification shared by many bioactive peptides. We analyzed peptides secreted from human medullary thyroid carcinoma TT cells that produce amidated peptides, and we identified two novel amidated peptides, designated neuroendocrine regulatory peptide (NERP)-1 and NERP-2. NERPs are derived from distinct regions of the neurosecretory protein that was originally identified as a product of a nerve growth factor-responsive gene in PC12 cells. Mass spectrometric analysis of the immunoprecipitate using specific antibodies as well as reversed phase-high performance liquid chromatography coupled with radioimmunoassay analysis of brain extract demonstrated the endogenous presence of NERP-1 and NERP-2 in the rat. NERPs are abundant in the paraventricular and supraoptic nuclei of the rat hypothalamus and colocalized frequently with vasopressin but rarely with oxytocin. NERPs dose-dependently suppressed vasopressin release induced by intracerebroventricular injection of hypertonic NaCl or angiotensin II in vivo. NERPs also suppressed basal and angiotensin II-induced vasopressin secretion from hypothalamic explants in vitro. Bioactivity of NERPs required C-terminal amidation. Anti-NERP IgGs canceled plasma vasopressin reduction in response to water loading, indicating that NERPs could be potent endogenous suppressors of vasopressin release. These findings suggest that NERPs are novel modulators in body fluid homeostasis.     

In vivo digestomics of milk proteins in human milk and infant formula using a suckling rat pup model

Human milk is the optimal mode of infant feeding for the first several months of life, and infant formulas serve as an alternative when breast-feeding is not possible. Milk proteins have a balanced amino acid composition and some of them provide beneficial bioactivities in their intact forms. They also encrypt a variety of bioactive peptides, possibly contributing to infant health and growth. However, there is limited knowledge of how milk proteins are digested in the gastrointestinal tract and bioactive peptides are released in infants. A peptidomic analysis was conducted to identify peptides released from milk proteins in human milk and infant formula, using a suckling rat pup model. Among the major milk proteins targeted, α-lactalbumin and β-casein in human milk, and β-lactoglobulin and β-casein in infant formula were the main sources of peptides, and these peptides covered large parts of the parental proteins’ sequences. Release of peptides was concentrated to specific regions, such as residues 70–92 of β-casein in human milk, residues 39–55 of β-lactoglobulin in infant formula, and residues 57–96 and 145–161 of β-CN in infant formula, where resistance to gastrointestinal digestion was suggested. In the context of bioactive peptides, release of fragments containing known bioactive peptides was confirmed, such as β-CN-derived opioid and antihypertensive peptides. It is therefore likely that these fragments are of biological significance in neonatal health and development.     

Transport of somatostatin and substance P by human P-glycoprotein

P-glycoprotein is an efflux pump for a broad spectrum of hydrophobic agents. We found that bioactive peptides including somatostatin and substance P inhibit ATP-dependent vincristine binding to P-glycoprotein-overexpressing K562/ADM membrane vesicles. Some of these bioactive peptides including somatostatin stimulate basal ATPase activity of P-glycoprotein; in contrast, other peptides including substance P inhibit it. The K562/ADM membrane vesicles showed an ATP-dependent, osmotically sensitive uptake of somatostatin and substance P, which was inhibited by valspodar, an inhibitor of P-glycoprotein. These findings suggested that certain bioactive peptides such as somatostatin and substance P directly interact with human P-glycoprotein as endogenous substrates for P-glycoprotein-mediated transport.