Characterization of the human submandibular/sublingual saliva glycoproteome using lectin affinity chromatography coupled to multidimensional protein identification technology

In-depth analysis of the salivary proteome is fundamental to understanding the functions of salivary proteins in the oral cavity and to reveal disease biomarkers involved in different pathophysiological conditions, with the ultimate goal of improving patient diagnosis and prognosis. Submandibular and sublingual glands contribute saliva rich in glycoproteins to the total saliva output, making them valuable sources for glycoproteomic analysis. Lectin-affinity chromatography coupled to mass spectrometry-based shotgun proteomics was used to explore the submandibular/sublingual (SM/SL) saliva glycoproteome. A total of 262 N- and O-linked glycoproteins were identified by multidimensional protein identification technology (MudPIT). Only 38 were previously described in SM and SL salivas from the human salivary N-linked glycoproteome, while 224 were unique. Further comparison analysis with SM/SL saliva of the human saliva proteome, revealed 125 glycoproteins not formerly reported in this secretion. KEGG pathway analyses demonstrated that many of these glycoproteins are involved in processes such as complement and coagulation cascades, cell communication, glycosphingolipid biosynthesis neo-lactoseries, O-glycan biosynthesis, glycan structures-biosynthesis 2, starch and sucrose metabolism, peptidoglycan biosynthesis or others pathways. In summary, lectin-affinity chromatography coupled to MudPIT mass spectrometry identified many novel glycoproteins in SM/SL saliva. These new additions to the salivary proteome may prove to be a critical step for providing reliable biomarkers in the diagnosis of a myriad of oral and systemic diseases.     

The scientific exploration of saliva in the post-proteomic era: from database back to basic function

The proteome of human saliva can be considered as being essentially completed. Diagnostic markers for a number of diseases have been identified among salivary proteins and peptides, taking advantage of saliva as an easy-to-obtain biological fluid. Yet, the majority of disease markers identified so far are serum components and not intrinsic proteins produced by the salivary glands. Furthermore, despite the fact that saliva is essential for protecting the oral integuments and dentition, little progress has been made in finding risk predictors in the salivary proteome for dental caries or periodontal disease. Since salivary proteins, and in particular the attached glycans, play an important role in interactions with the microbial world, the salivary glycoproteome and other post-translational modifications of salivary proteins need to be studied. Risk markers for microbial diseases, including dental caries, are likely to be discovered among the highly glycosylated major protein species in saliva. This review will attempt to raise new ideas and also point to under-researched areas that may hold promise for future applicability in oral diagnostics and prediction of oral disease.     

黄曲霉菌次级代谢的组学研究进展

黄曲霉菌Aspergillusflavus是一种好氧型腐生真菌,其次级代谢产物黄曲霉毒素主要由黄曲霉和寄生曲霉产生,对诸如玉米、花生等在内的农作物侵染已经严重危及食品安全以及人和动物的健康。近年来,不同组学研究发展迅速,生物学研究的基础数据平台逐步建立。从基因组到转录组、蛋白质组、代谢组等组学技术在对黄曲霉菌次级代谢相关的研究中已有较多应用。本文概述了基因组、转录组、蛋白质组以及代谢组等组学技术在黄曲霉次级代谢研究中所取得的重要进展,并提出了相关研究的发展趋势以及有待解决的问题。为深入了解黄曲霉的生物学功能提供了重要的线索,并为今后的研究奠定了坚实的基础。     

Characteristics of human saliva proteome and peptidome

Recent studies on the characteristics of saliva proteome and peptidome greatly expanded our understanding of this biological fluid. Athough many scientists consider saliva to be an ideal biosubstrate in diagnosis of the human body state; currently, the research in this area is at the data accumulation stage. The physiology of saliva and salivary glands, as well as characteristics of interaction between the saliva proteins and the oral cavity microorganisms, has been insufficiently studied yet. The lack of standardization in collecting the saliva samples and in the proteome research protocols, and the requirements for sample representativeness introduce discrepancies in the results obtained by different researchers. Addressing these problems will allow the wide use of saliva proteome as a complex indicator of the functional state of the human body.     

宏基因组学及其在口腔微生物研究中的应用

宏基因组是指环境中所有微生物的遗传物质总和。宏基因组学技术可以最大限度地利用环境中的微生物资源,受到了国内外微生物研究者的重点关注。口腔中寄居着大量的微生物群落,以往对口腔疾病微生物的研究大多局限于单纯的细菌培养技术,然而,由于培养技术的局限性,部分微生物很难或根本不能培养,宏基因组学技术打破了这一局限性,帮助人类发掘更丰富的口腔微生物资源。最近,以宏基因组学测序为基础的研究描绘出了口腔生态系统的图谱,越来越多的实验证明口腔微生物组在各种口腔疾病甚至全身系统性疾病中的重要作用。同时,这也为基于人类微生物组的诊断和治疗开辟了新的途径。本综述旨在说明宏基因组学是研究人类口腔疾病及全身疾病相关微生物的得力工具,而且具有广阔的发展前景,同时也讨论了宏基因组学在应用中有待克服的局限性。     

2014蛋白质组学专刊序言

蛋白质组学研究是后基因组学时代最重要的功能基因组学研究之一,与医学生物学、化学、物理学、信息学以及现代技术等关系十分密切。为了检阅近年来国内外蛋白质组学某些重要研究进展,探索其可能的应用范围,讨论其存在的问题,展望其发展前景,特组织出版"蛋白质组学专刊"。本期专刊包括综述和研究论文两部分,内容主要涉及不同物种(包括人类、哺乳类动物、原核生物、放线菌等)蛋白质组学研究、蛋白质组学重要方法学与技术研究(包括串联质谱分析、尿蛋白膜保存法、定量蛋白质组学分折、meta分析等)和蛋白质组功能与应用研究(包括蜘蛛毒素蛋白质组、磷酸化蛋白质组、卵母细胞和早期胚胎蛋白质组、肝脏纤维化蛋白质组、分枝杆菌耐药的蛋白质组等)。     

功能基因组学及其研究方法

随着水稻基因组测序工作的完成以及人类基因组计划(HGP)的顺利进行,生命科学的研究已经进入了后基因组时代。对功能基因组学的研究方法进行了综述,主要包括:微阵列、基因表达系列分析、反义RNA和RNAi、基因敲除、基因陷阱、蛋白质组、生物信息学和功能基因组系统学等。     

Saliva proteome research: current status and future outlook

Human saliva harbours proteins of clinical relevance and about 30% of blood proteins are also present in saliva. This highlights that saliva can be used for clinical applications just as urine or blood. However, the translation of salivary biomarker discoveries into clinical settings is hampered by the dynamics and complexity of the salivary proteome. This review focuses on the current status of technological developments and achievements relating to approaches for unravelling the human salivary proteome. We discuss the dynamics of the salivary proteome, as well as the importance of sample preparation and processing techniques and their influence on downstream protein applications; post-translational modifications of salivary proteome and protein: protein interactions. In addition, we describe possible enrichment strategies for discerning post-translational modifications of salivary proteins, the potential utility of selected-reaction-monitoring techniques for biomarker discovery and validation, limitations to proteomics and the biomarker challenge and future perspectives. In summary, we provide recommendations for practical saliva sampling, processing and storage conditions to increase the quality of future studies in an emerging field of saliva clinical proteomics. We propose that the advent of technologies allowing sensitive and high throughput proteome-wide analyses, coupled to well-controlled study design, will allow saliva to enter clinical practice as an alternative to blood-based methods due to its simplistic nature of sampling, non-invasiveness, easy of collection and multiple collections by untrained professionals and cost-effective advantages.     

Large-scale identification of proteins in human salivary proteome by liquid chromatography/mass spectrometry and two-dimensional gel electrophoresis-mass spectrometry

Human saliva contains a large number of proteins and peptides (salivary proteome) that help maintain homeostasis in the oral cavity. Global analysis of human salivary proteome is important for understanding oral health and disease pathogenesis. In this study, large-scale identification of salivary proteins was demonstrated by using shotgun proteomics and two-dimensinal gel electrophoresis-mass spectrometry (2-DE-MS). For the shotgun approach, whole saliva proteins were prefractionated according to molecular weight. The smallest fraction, presumably containing salivary peptides, was directly separated by capillary liquid chromatography (LC). However, the large protein fractions were digested into peptides for subsequent LC separation. Separated peptides were analyzed by on-line electrospray tandem mass spectrometry (MS/MS) using a quadrupole-time of flight mass spectrometer, and the obtained spectra were automatically processed to search human protein sequence database for protein identification. Additionally, 2-DE was used to map out the proteins in whole saliva. Protein spots 105 in number were excised and in-gel digested; and the resulting peptide fragments were measured by matrix-assisted laser desorption/ionization-mass spectrometry and sequenced by LC-MS/MS for protein identification. In total, we cataloged 309 proteins from human whole saliva by using these two proteomic approaches.     

细菌群体感应的蛋白质组学研究进展

细菌群体感应是指细菌能合成、释放和感应一些类激素小分子信号,从而调控群体行为并对其作出应答反应。介导细菌群体感应的信号分子有多种,它们参与调节细菌许多重要生物学功能。目前对此研究的主要手段是基因组学和转录组学。然而近年来,基因组测序技术的不断发展为另一种新兴方法——以比较和功能性为基础的蛋白质组学法奠定了基础。所不同的是,传统方法只能局限性研究某些基因或蛋白,而蛋白质组学法能检测出生物体基因表达的全部蛋白,它也因此逐渐受到人们的广泛关注。主要从研究较多的三类信号分子方面描述如何利用蛋白盾组举法解析细菌交流的“语言”。