{"title":"Purification of Proteins: Between Meaning and Different Methods","authors":"H. Amer","doi":"10.5772/intechopen.86587","DOIUrl":"https://doi.org/10.5772/intechopen.86587","url":null,"abstract":"Purification of protein attracts the scientists’ attention toward science in 1926, as Somnar started purification and crystallization of urease from yeast. As years go forward, protein purification strategies updated from using dextrose passing through DEAE-cellulose and ended by affinity chromatography. In this chapter we will describe some of the concepts and the differences between traditional and novel purification methods, in order to point out how a researcher can start the protein purification techniques. Different fundamental purification steps from plant sample will be discussed in this chapter including isolation, concentration, ion exchanger, and affinity chromatography, as well as the important additives that a researcher should add in order to gain a high purification fold.","PeriodicalId":399676,"journal":{"name":"Proteomics Technologies and Applications","volume":"22 1","pages":"0"},"PeriodicalIF":0.0,"publicationDate":"2019-09-02","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"125042593","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
{"title":"2D Gel Electrophoresis to Address Biological Issues","authors":"D. Scumaci, G. Cuda","doi":"10.5772/INTECHOPEN.86211","DOIUrl":"https://doi.org/10.5772/INTECHOPEN.86211","url":null,"abstract":"Two-dimensional (2D) gel electrophoresis is a high-resolution technique for the study of proteome. This chapter describes how it can be applied to characterize specific differences in the proteome profile of breast cancer cells following gene target interference. The proteome is the complete set of proteins encoded by a genome, and proteomic analysis consists in profiling the whole proteins expressed in a given cell, tissue, organ, or organism. Proteomic expression has the main purpose of qualitatively and quantitatively comparing proteins expressed under physiological and/or pathological conditions. Although it is not the unique approach used in modern proteomics, two-dimensional electrophoresis (2DE) is unrivaled allowing simultaneous separation of thousands of proteins and the detection of posttranslational modification, not predictable through genome analysis. 2DE combines two physical principles to separate complex protein mixtures: the isoelectric point and the molecular weight. The result is a gel map in which each protein isoform present in the sample can be visualized as a spot, analyzed, quantified, and identified by mass spectrometry analysis. Here we outline features and advantages of the 2DE-based proteomic approach and we describe how 2DE meets biochemistry and molecular biology to address specific issues. enhanced cisplatin cytotoxicity in tumor cells having both BRCA network and TP53 disruptions.","PeriodicalId":399676,"journal":{"name":"Proteomics Technologies and Applications","volume":"19 1","pages":"0"},"PeriodicalIF":0.0,"publicationDate":"2019-05-31","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"125851282","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
S. Ponnikorn, Siripath Peter Kong, Sasipim Thitivirachawat, Chanawin Tanjasiri, S. Tungpradabkul, S. Hongeng
{"title":"Proteomic Analysis of β-Thalassemia/HbE: A Perspective from Hematopoietic Stem Cells (HSCs)","authors":"S. Ponnikorn, Siripath Peter Kong, Sasipim Thitivirachawat, Chanawin Tanjasiri, S. Tungpradabkul, S. Hongeng","doi":"10.5772/INTECHOPEN.85863","DOIUrl":"https://doi.org/10.5772/INTECHOPEN.85863","url":null,"abstract":"β-Thalassemia/HbE is highly prevalent in Southeast Asian countries, especially Thailand. It is a severe hereditary anemia disease involving ineffective erythropoiesis in the bone marrow and peripheral tissues. The excess of alpha globin and iron overload contribute to elevated oxidative damages leading to a premature cell death of erythroid cells and a diminished terminal differentiation of reticulocytes. Although proteomic approach would gain a comprehensive picture of the complex pathophysiology of human bone marrow and hematopoietic stem cells (HSCs), obtaining sufficient clinical specimens remains an important issue. The employment of mass spectrometry (MS)-based proteomic profiling could overcome these constrains and provide useful insights into the cellular constituents and microenvironment in bone marrow milieu. In this chapter, we summarize the comparative proteomic studies analyzing CD34+/HSCs and bone marrow niche proteins. Under ineffective erythropoiesis, in-depth analyses of various proteome profiles revealed many of which have putative functions. Importantly, dysregulated cell death and survival signaling pathways could explain the deleterious pathogenesis of β-thalassemia/HbE.","PeriodicalId":399676,"journal":{"name":"Proteomics Technologies and Applications","volume":"1 1","pages":"0"},"PeriodicalIF":0.0,"publicationDate":"2019-04-19","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"129474968","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
R. Albulescu, A. Petrescu, M. Sarbu, Alice Grigore, Raluca Ica, C. Munteanu, A. Albulescu, Ioana V. Militaru, A. Zamfir, S. Petrescu, C. Tanase
{"title":"Mass Spectrometry for Cancer Biomarkers","authors":"R. Albulescu, A. Petrescu, M. Sarbu, Alice Grigore, Raluca Ica, C. Munteanu, A. Albulescu, Ioana V. Militaru, A. Zamfir, S. Petrescu, C. Tanase","doi":"10.5772/INTECHOPEN.85609","DOIUrl":"https://doi.org/10.5772/INTECHOPEN.85609","url":null,"abstract":"Cancer is considered as the second cause of morbidity and also mortality, after cardiovascular diseases. Despite the immense progress in efficacious biomarkers made in the present time, there are very few of them that can timely detect cancers or that can predict treatment outcomes or stratify patients according to their response to the treatment. Among other modern instruments involved in the research of this disease, proteomics emerged strongly, since it analyzes the “molec-ular effectors.” Although it has some setbacks (like the lack of amplification of the signal), it is however one of the best means of investigating the presence and causes and predicting the evolution patterns of the disease. This chapter describes briefly pre-analytical (pre-MS steps), the main concepts, and the MS equipment used for such applications, followed by the presentation of several proteomic applications in melanoma, glioblastoma, pancreatic cancer, and colon cancer. the differential peptide display mass spectrometric signal intensity of the peptides in CSF Spearman’s rank the peptidomes. N distinguished controls in all applied individu-als. The results identified nine IgG N-glycans that were differentially expressed in patient groups. Moreover, five of them were significantly modified in CRC patients in all TNM stages. Principal component analysis (PCA) indicated evident differentiation between benign, cancer patients and normal individuals. The ROC analysis suggests that these five IgG N-glycans were correlated with the progression of","PeriodicalId":399676,"journal":{"name":"Proteomics Technologies and Applications","volume":"108 1 1","pages":"0"},"PeriodicalIF":0.0,"publicationDate":"2019-04-12","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"131900126","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
{"title":"Invasiveness-Related Proteomic Variations and Molecular Network Changes in Human Nonfunctional Pituitary Adenomas","authors":"X. Zhan, X. Zhan, Xiaowei Wang","doi":"10.5772/INTECHOPEN.85546","DOIUrl":"https://doi.org/10.5772/INTECHOPEN.85546","url":null,"abstract":"The invasive characteristic of nonfunctional pituitary adenoma (NFPA) is an important clinical problem without a clear molecular mechanism, which severely challenges its treatment strategy. Clarification of the proteomic alterations between invasive and non-invasive NFPAs is the key step for in-depth understanding of its mechanisms and discovering reliably invasive biomarkers. Two-dimensional gel electrophoresis (2DGE)-based comparative proteomics was carried out between four invasive and four non-invasive NFPAs. A total of 64 upregulated protein-spots and 39 downregulated protein-spots were identified among 24 (invasive n = 12; non-invasive n = 12) 2DGE maps (ca. 1200 spots/gel). Mass spectrometry identified 30 upregulated proteins and 27 downregulated proteins between invasive and noninvasive NFPAs. Those 57 differentially expressed proteins are involved in multiple biological functions, including oxidative stress, mitochondrial dysfunction, MAPK signaling alteration, proteolysis abnormality, CDK-C signaling, amyloid processing, and TR/RXR activation. These findings provide important clues to insights into molecular mechanisms of invasive NFPAs and to discovery of effective biomarkers for effective treatment of invasive NFPA patients.","PeriodicalId":399676,"journal":{"name":"Proteomics Technologies and Applications","volume":"104 1","pages":"0"},"PeriodicalIF":0.0,"publicationDate":"2019-04-10","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"131577995","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
{"title":"Precipitation of Detergent-Containing Samples for Top-Down and Bottom-Up Proteomics","authors":"A. Doucette, Andrew M. J. Crowell","doi":"10.5772/INTECHOPEN.85547","DOIUrl":"https://doi.org/10.5772/INTECHOPEN.85547","url":null,"abstract":"Prior to proteome analysis by mass spectrometry (MS), protein mixtures must first be subject to various sample preparation steps. The goal is to isolate proteins in high yield, and with high purity. Liquid chromatography (LC) separation is also integral to comprehensive proteome characterization, and so a key component of sample preparation is simply to solubilize the proteome in LC-MS compatible solvents. Hydrophobic proteins (membrane proteins) represent a greater chal-lenge to maintain protein solubility during sample preparation. Sodium dodecyl sulfate (SDS) is a favored detergent to solubilize proteins, and also is used to impart mass-based fractionation (i.e., SDS PAGE, GELFrEE). However, SDS is incompat-ible with downstream LC-MS analysis. Fortunately, effective strategies for SDS removal do exist, which permits the use of this surfactant in proteomics workflows. Here we highlight an approach that is grounded in the classic technique—protein precipitation. The technique has been updated and has recently seen a revival as a strategy permitting high protein recovery, with exceptional purity. Moreover, with aid of simple disposable spin cartridges, protein precipitation can meet the needs of high throughput, automated, and reproducible proteome purification, enabling the analysis of SDS-containing samples in both top-down and bottom-up formats.","PeriodicalId":399676,"journal":{"name":"Proteomics Technologies and Applications","volume":"158 1","pages":"0"},"PeriodicalIF":0.0,"publicationDate":"2019-04-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"133482232","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
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