Pub Date : 2018-09-12DOI: 10.5772/INTECHOPEN.76889
X. Zhan, Na Li
Protein tyrosine nitration is an important molecular event in nervous system tumor such as glioma and pituitary adenomas. It is the essential step to identify the protein targets and exact modified sites of tyrosine nitration for addressing the biological roles of pro - tein tyrosine nitration in nervous system tumors and discovering effective biomarkers to understand in-depth molecular mechanisms and determine new diagnosis strategy and novel therapeutic targets. One/two-dimensional gel electrophoresis (1DGE, 2DGE), or nitrotyrosine affinity column (NTAC), coupled with tandem mass spectrometry (MS/MS) have been successfully applied in the analysis of nitroproteins in nervous system tumors. This article address the basic concept of protein tyrosine nitration, nitroproteomics meth - odology based on gel electrophoresis/immunoaffinity enrichment and tandem mass spectrometry, and the current status of nitroprotein study in nervous system tumors. The established nitroproteomics approach is easily translated to study other diseases. NO and RNS are important inflammatory mediators [ 15 ]; and (3) increased production of NO, peroxinitrite and superoxide, occurs in nervous system tumors [ 16 ]; (4) higher levels of nitrotyrosine are observed in nervous system tumors than normal tissues with biochemical approaches and immunohistochemical, and only protein nitrotubulin and protein nitro-p53 have been determined in human nervous system tumors [ 17]. Furthermore, the amino acid analog 3-nitrotyrosine due to functional and morphological injury of mouse-neuroblastoma cell lines and rat-glioma cell lines [ 18 ]. These studies demonstrated the importance of protein tyrosine nitration in the pathogenesis of nervous system tumors. Illustrating the functions of nitroproteins might reveal in-depth molecular mechanisms and biological function of tyro sine nitration in human nervous system tumors. Literature-based review and comprehen sive annotation of proteins on the SwissProt website were used to expound the nitroprotein domains/motifs, location of nitrotyrosine sites and possible signaling pathways relevant to nervous system tumors. Nitroproteins took part in multiple biological processes in the devel opment of tumors as follows: (a) tumor cell migration and invasion [ 19 ]; (b) cell proliferation and apoptosis [20]; (c) chemotherapy resistance [ 21 ]; (d) signal transduction [ 22]; (e) phenotypic dedifferentiation [23]; (f) microtubule dynamic stabilization [24 ]; (g) tumor recurrence; (h) others such as immunoreaction and post-transcriptional regulation. Moreover, the discov ery of tyrosine nitration being a reversible reaction [ 25 ] and having a competition between phosphorylation motif [ 26], led us to speculate that dynamic process of protein nitration might also be regulated and controlled. However, no definite target of intervention is found for tyrosine nitration in human nervous system tumors. It takes long time to study tumor-related nitroproteins and t
{"title":"The Use of Gel Electrophoresis and Mass Spectrometry to Identify Nitroproteins in Nervous System Tumors","authors":"X. Zhan, Na Li","doi":"10.5772/INTECHOPEN.76889","DOIUrl":"https://doi.org/10.5772/INTECHOPEN.76889","url":null,"abstract":"Protein tyrosine nitration is an important molecular event in nervous system tumor such as glioma and pituitary adenomas. It is the essential step to identify the protein targets and exact modified sites of tyrosine nitration for addressing the biological roles of pro - tein tyrosine nitration in nervous system tumors and discovering effective biomarkers to understand in-depth molecular mechanisms and determine new diagnosis strategy and novel therapeutic targets. One/two-dimensional gel electrophoresis (1DGE, 2DGE), or nitrotyrosine affinity column (NTAC), coupled with tandem mass spectrometry (MS/MS) have been successfully applied in the analysis of nitroproteins in nervous system tumors. This article address the basic concept of protein tyrosine nitration, nitroproteomics meth - odology based on gel electrophoresis/immunoaffinity enrichment and tandem mass spectrometry, and the current status of nitroprotein study in nervous system tumors. The established nitroproteomics approach is easily translated to study other diseases. NO and RNS are important inflammatory mediators [ 15 ]; and (3) increased production of NO, peroxinitrite and superoxide, occurs in nervous system tumors [ 16 ]; (4) higher levels of nitrotyrosine are observed in nervous system tumors than normal tissues with biochemical approaches and immunohistochemical, and only protein nitrotubulin and protein nitro-p53 have been determined in human nervous system tumors [ 17]. Furthermore, the amino acid analog 3-nitrotyrosine due to functional and morphological injury of mouse-neuroblastoma cell lines and rat-glioma cell lines [ 18 ]. These studies demonstrated the importance of protein tyrosine nitration in the pathogenesis of nervous system tumors. Illustrating the functions of nitroproteins might reveal in-depth molecular mechanisms and biological function of tyro sine nitration in human nervous system tumors. Literature-based review and comprehen sive annotation of proteins on the SwissProt website were used to expound the nitroprotein domains/motifs, location of nitrotyrosine sites and possible signaling pathways relevant to nervous system tumors. Nitroproteins took part in multiple biological processes in the devel opment of tumors as follows: (a) tumor cell migration and invasion [ 19 ]; (b) cell proliferation and apoptosis [20]; (c) chemotherapy resistance [ 21 ]; (d) signal transduction [ 22]; (e) phenotypic dedifferentiation [23]; (f) microtubule dynamic stabilization [24 ]; (g) tumor recurrence; (h) others such as immunoreaction and post-transcriptional regulation. Moreover, the discov ery of tyrosine nitration being a reversible reaction [ 25 ] and having a competition between phosphorylation motif [ 26], led us to speculate that dynamic process of protein nitration might also be regulated and controlled. However, no definite target of intervention is found for tyrosine nitration in human nervous system tumors. It takes long time to study tumor-related nitroproteins and t","PeriodicalId":186044,"journal":{"name":"Electrophoresis - Life Sciences Practical Applications","volume":"1 1","pages":"0"},"PeriodicalIF":0.0,"publicationDate":"2018-09-12","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"116230214","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}
Pub Date : 2018-09-12DOI: 10.5772/INTECHOPEN.75782
Koubaa Anouar, Bargui Mansour
This chapter deals with the study of the elaboration of a stable suspension of TiO2 nanoparticles and their incorporation by electrophoretic deposition into pores of anodized 5754 aluminum alloy. The as-synthesized TiO2 nanopowder was characterized by X-ray diffraction, scanning and transmission electron microscopy (TEM), and infrared spectroscopy. During this work, transmission electronic microscopy (TEM) analysis showed that the resulting particles had a narrow size distribution with crystallite size of about 15 nm. The zeta potential and stability of TiO2 nanoparticles dispersed with poly(acrylic acid) (PAA) in aqueous solution were also measured. Porous anodic film was elaborated in phosphoric acid electrolyte and then filled by TiO2 particles using electrophoresis method. Furthermore, the effect of PAA content and pH on the suspension stability has been investigated. It was also demonstrated that buffered suspension by adding glycine avoids gelating phenomena which inhibits the insertion of nanoparticles inside the pores of anodic film. It was noted also that the electric field already applied greatly influences the electrophoretic deposition process (EPD). FEG-SEM observations showed that larger (125 nm diameter) and linear pores of 6 μm in length are successfully filled in 5 min. Finally, the composite anodic film tribological behavior was studied and the obtained results revealed that the insertion of the TiO2 nanoparticles into the pores of the anodic film improves its tribological properties.
{"title":"Improving Tribological Behavior of Porous Anodic Film by Electrophoretic Impregnation by a Tio2 Synthesized Nanoparticle","authors":"Koubaa Anouar, Bargui Mansour","doi":"10.5772/INTECHOPEN.75782","DOIUrl":"https://doi.org/10.5772/INTECHOPEN.75782","url":null,"abstract":"This chapter deals with the study of the elaboration of a stable suspension of TiO2 nanoparticles and their incorporation by electrophoretic deposition into pores of anodized 5754 aluminum alloy. The as-synthesized TiO2 nanopowder was characterized by X-ray diffraction, scanning and transmission electron microscopy (TEM), and infrared spectroscopy. During this work, transmission electronic microscopy (TEM) analysis showed that the resulting particles had a narrow size distribution with crystallite size of about 15 nm. The zeta potential and stability of TiO2 nanoparticles dispersed with poly(acrylic acid) (PAA) in aqueous solution were also measured. Porous anodic film was elaborated in phosphoric acid electrolyte and then filled by TiO2 particles using electrophoresis method. Furthermore, the effect of PAA content and pH on the suspension stability has been investigated. It was also demonstrated that buffered suspension by adding glycine avoids gelating phenomena which inhibits the insertion of nanoparticles inside the pores of anodic film. It was noted also that the electric field already applied greatly influences the electrophoretic deposition process (EPD). FEG-SEM observations showed that larger (125 nm diameter) and linear pores of 6 μm in length are successfully filled in 5 min. Finally, the composite anodic film tribological behavior was studied and the obtained results revealed that the insertion of the TiO2 nanoparticles into the pores of the anodic film improves its tribological properties.","PeriodicalId":186044,"journal":{"name":"Electrophoresis - Life Sciences Practical Applications","volume":"87 1","pages":"0"},"PeriodicalIF":0.0,"publicationDate":"2018-09-12","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"124016409","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}
Pub Date : 2018-09-12DOI: 10.5772/INTECHOPEN.75902
I. Vikhlyantsev, Z. Podlubnaya
Titin (also known as connectin) is a giant elastic protein of striated and smooth muscles of vertebrates. The molecular weight of its isoforms is 3.0–3.7 MDa in striated muscles and 0.5–2.0 MDa in smooth muscles. Titin was discovered 40 years ago using the sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE). At the present time, this method has not lost its relevance but has undergone a number of modifications that improve visualization of giant titin isoforms in the gel. This chapter provides historical insights into the technical aspects of the electrophoresis methods used to identify titin and its isoforms. We focus on the peculiarities of the technique because of which titin molecules remain intact and its high molecular weight isoforms can be visualized. Electrophoretic testing of changes in titin content in muscles can be used in medical practice to diagnose pathological processes and evaluate effective approaches to their correction. was performed in vertical agarose- strengthened 2.1% polyacrylamide gel (8 × 10 × 0.1 cm). (1) m. soleus (control); (2) m. soleus (proteolysis, 1 h); (3) left ventricle of heart (control); (4) left ventricle of heart (proteolysis, 30 min). Proteolytic cleavage of titin was performed under the influence of endogenous muscular proteases. To this end, small pieces of muscle tissue (20–30 mg) were held for 30–60 min at 25–30°C. Then, 2–3 mg pieces were taken from the muscle sample and placed into solubilizing solution (10 mM Tris–HCl, 1.2% SDS, 10% glycerol, 2% β-mercaptoethanol or 75 mM DTT, 8–10 μg/ml of leupeptin or E64, pH 7.0) for the extraction and further electrophoretic testing of the proteins. T3300 is probably the proteolytic fragment of NT titin with molecular weight of ~3300 kDa.
连接蛋白(又称连接蛋白)是脊椎动物横纹肌和平滑肌中的一种巨大的弹性蛋白。其异构体在横纹肌中分子量为3.0 ~ 3.7 MDa，在平滑肌中分子量为0.5 ~ 2.0 MDa。Titin是40年前用十二烷基硫酸钠-聚丙烯酰胺凝胶电泳(SDS-PAGE)发现的。目前，这种方法并没有失去它的相关性，但已经经历了一些修改，提高了凝胶中巨大的titin异构体的可视化。本章提供了历史的见解到电泳方法的技术方面，用于鉴定titin和它的同种异构体。我们专注于该技术的特殊性，因为它的titin分子保持完整，其高分子量的同工异构体可以可视化。肌肉中肌球蛋白含量变化的电泳检测可用于医学实践中诊断病理过程和评估其纠正的有效方法。在琼脂糖增强的2.1%聚丙烯酰胺垂直凝胶(8 × 10 × 0.1 cm)中进行。(1)比目鱼(对照);(2) m. soleus(蛋白水解，1小时);(3)左心室(对照);(4)左心室(蛋白水解，30分钟)。在内源性肌肉蛋白酶的作用下，titin蛋白水解裂解。为此，将小块肌肉组织(20-30 mg)在25-30°C下保存30-60分钟。然后取2-3 mg肌肉样品，置于增溶液(10 mM Tris-HCl, 1.2% SDS, 10%甘油，2% β-巯基乙醇或75 mM DTT, 8-10 μg/ml胰肽或E64, pH 7.0)中提取并进一步电泳检测蛋白质。T3300可能是NT titin的蛋白水解片段，分子量约为3300 kDa。
{"title":"Peculiarities of SDS-PAGE of Titin/Connectin","authors":"I. Vikhlyantsev, Z. Podlubnaya","doi":"10.5772/INTECHOPEN.75902","DOIUrl":"https://doi.org/10.5772/INTECHOPEN.75902","url":null,"abstract":"Titin (also known as connectin) is a giant elastic protein of striated and smooth muscles of vertebrates. The molecular weight of its isoforms is 3.0–3.7 MDa in striated muscles and 0.5–2.0 MDa in smooth muscles. Titin was discovered 40 years ago using the sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE). At the present time, this method has not lost its relevance but has undergone a number of modifications that improve visualization of giant titin isoforms in the gel. This chapter provides historical insights into the technical aspects of the electrophoresis methods used to identify titin and its isoforms. We focus on the peculiarities of the technique because of which titin molecules remain intact and its high molecular weight isoforms can be visualized. Electrophoretic testing of changes in titin content in muscles can be used in medical practice to diagnose pathological processes and evaluate effective approaches to their correction. was performed in vertical agarose- strengthened 2.1% polyacrylamide gel (8 × 10 × 0.1 cm). (1) m. soleus (control); (2) m. soleus (proteolysis, 1 h); (3) left ventricle of heart (control); (4) left ventricle of heart (proteolysis, 30 min). Proteolytic cleavage of titin was performed under the influence of endogenous muscular proteases. To this end, small pieces of muscle tissue (20–30 mg) were held for 30–60 min at 25–30°C. Then, 2–3 mg pieces were taken from the muscle sample and placed into solubilizing solution (10 mM Tris–HCl, 1.2% SDS, 10% glycerol, 2% β-mercaptoethanol or 75 mM DTT, 8–10 μg/ml of leupeptin or E64, pH 7.0) for the extraction and further electrophoretic testing of the proteins. T3300 is probably the proteolytic fragment of NT titin with molecular weight of ~3300 kDa.","PeriodicalId":186044,"journal":{"name":"Electrophoresis - Life Sciences Practical Applications","volume":"27 1","pages":"0"},"PeriodicalIF":0.0,"publicationDate":"2018-09-12","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"121754225","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}
Pub Date : 2018-09-12DOI: 10.5772/INTECHOPEN.76880
G. Brunborg, L. Rolstadaas, K. Gutzkow
The comet assay is a sensitive technique to measure lesions in DNA, based on electropho- retic separation of DNA from cells embedded in agarose. Movement of DNA fragments is determined by the potential (V/cm), the time, and the viscosity of the medium (agarose). There is historically considerable confusion as to other factors, that is, current, liquid depths, circulation of the liquid, and temperature. Lack of standardization of electropho- resis including suboptimal power supplies and electrophoresis tanks causes considerable variations within and between laboratories. Ring trials have not been able to clearly identify the cause(s) of variation. Comparison of comet data from cohorts of human blood lymphocytes is used in the COST project hCOMET to identify early biomarkers of the disease. This calls for standardization of analysis. We performed measurements of electric potentials in a tank using multiple electrodes. Variations (time/position) were reduced by circulating electrophoresis liquid at 10% (volume) per min; this also stabilized the temperature. Circulation was accompanied by only slightly reduced variation in DNA damage among 384 irradiated cell samples electrophoresed concomitantly. In conclusion, comparing data between laboratories and cohorts must give emphasis to electrophoresis condi- tions. Results should be specified with respect to voltage (V/cm), time, and agarose concentration. We expect that suitable correction factors for these parameters may reduce inter-laboratory variations in comet data, allowing more precise comparison of results from different human cohorts.
{"title":"Electrophoresis in the Comet Assay","authors":"G. Brunborg, L. Rolstadaas, K. Gutzkow","doi":"10.5772/INTECHOPEN.76880","DOIUrl":"https://doi.org/10.5772/INTECHOPEN.76880","url":null,"abstract":"The comet assay is a sensitive technique to measure lesions in DNA, based on electropho- retic separation of DNA from cells embedded in agarose. Movement of DNA fragments is determined by the potential (V/cm), the time, and the viscosity of the medium (agarose). There is historically considerable confusion as to other factors, that is, current, liquid depths, circulation of the liquid, and temperature. Lack of standardization of electropho- resis including suboptimal power supplies and electrophoresis tanks causes considerable variations within and between laboratories. Ring trials have not been able to clearly identify the cause(s) of variation. Comparison of comet data from cohorts of human blood lymphocytes is used in the COST project hCOMET to identify early biomarkers of the disease. This calls for standardization of analysis. We performed measurements of electric potentials in a tank using multiple electrodes. Variations (time/position) were reduced by circulating electrophoresis liquid at 10% (volume) per min; this also stabilized the temperature. Circulation was accompanied by only slightly reduced variation in DNA damage among 384 irradiated cell samples electrophoresed concomitantly. In conclusion, comparing data between laboratories and cohorts must give emphasis to electrophoresis condi- tions. Results should be specified with respect to voltage (V/cm), time, and agarose concentration. We expect that suitable correction factors for these parameters may reduce inter-laboratory variations in comet data, allowing more precise comparison of results from different human cohorts.","PeriodicalId":186044,"journal":{"name":"Electrophoresis - Life Sciences Practical Applications","volume":"9 1","pages":"0"},"PeriodicalIF":0.0,"publicationDate":"2018-09-12","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"130936067","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}
Pub Date : 2018-09-12DOI: 10.5772/INTECHOPEN.76322
D. Heinová, Z. Kostecká, E. Petrovová
Lactate dehydrogenase (LDH) is a tetrameric enzyme that in vertebrates exists in five electrophoretically distinguishable forms known as isoenzymes. According to their different mobility to anode, they are denoted LDH 1 (H 4 ), LDH 2 (H 3 M), LDH 3 (H 2 M 2 ), LDH 4 (HM 3 ), and LDH 5 (M 4 ). A buffer system of the pH values 8.6–8.8 is commonly used for the separation of these isoenzymes in mammals. In the case of bird LDHs, the observation of five fractions is very difficult under this condition as they usually produce a pattern of one diffuse zone. Isoelectric focusing technique (IEF) in the pH range of 3–9 enabled a good and clear resolution of all five bird LDHs. Using this technique, it was also possible to observe the pattern in some tissues of chicken embryo. pattern where anodic LDH 1 -LDH 3 dominate over cathodic form/s. Mammalian tissues pattern of lactate dehydrogenase isoenzymes differs from species to species with the highest enzyme activity in the skeletal muscle followed by heart and liver. Chicken adult and embryonic lac tate dehydrogenases differ each other especially in the pattern of breast muscle with all five isoenzymes being present in the tissue of embryonic origin.
{"title":"Lactate Dehydrogenase Isoenzyme Electrophoretic Pattern in Serum and Tissues of Mammalian and Bird Origin","authors":"D. Heinová, Z. Kostecká, E. Petrovová","doi":"10.5772/INTECHOPEN.76322","DOIUrl":"https://doi.org/10.5772/INTECHOPEN.76322","url":null,"abstract":"Lactate dehydrogenase (LDH) is a tetrameric enzyme that in vertebrates exists in five electrophoretically distinguishable forms known as isoenzymes. According to their different mobility to anode, they are denoted LDH 1 (H 4 ), LDH 2 (H 3 M), LDH 3 (H 2 M 2 ), LDH 4 (HM 3 ), and LDH 5 (M 4 ). A buffer system of the pH values 8.6–8.8 is commonly used for the separation of these isoenzymes in mammals. In the case of bird LDHs, the observation of five fractions is very difficult under this condition as they usually produce a pattern of one diffuse zone. Isoelectric focusing technique (IEF) in the pH range of 3–9 enabled a good and clear resolution of all five bird LDHs. Using this technique, it was also possible to observe the pattern in some tissues of chicken embryo. pattern where anodic LDH 1 -LDH 3 dominate over cathodic form/s. Mammalian tissues pattern of lactate dehydrogenase isoenzymes differs from species to species with the highest enzyme activity in the skeletal muscle followed by heart and liver. Chicken adult and embryonic lac tate dehydrogenases differ each other especially in the pattern of breast muscle with all five isoenzymes being present in the tissue of embryonic origin.","PeriodicalId":186044,"journal":{"name":"Electrophoresis - Life Sciences Practical Applications","volume":"53 1","pages":"0"},"PeriodicalIF":0.0,"publicationDate":"2018-09-12","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"133195279","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}
Pub Date : 2018-09-12DOI: 10.5772/INTECHOPEN.75125
S. Naryzhny
The main intricacy in the human proteome is that it is tremendously complex and com- posed of diverse and heterogeneous gene products. These products are called protein species or proteoforms and are the smallest units of the proteome. In pursuit of the comprehensive profiling of the human proteome, significant advances should be performed. The approaches that allow disclosing and keeping the information about human proteome using two-dimensional gel electrophoresis (2DE) are described. Experimental identification methods such as mass spectrometry of high resolution and sensitiv- ity (MALDI-TOF MS and ESI LC-MS/MS) or immunodetection in combination with bioinformatics and 2DE can be used for the development of a comprehensive knowledge base of the human proteome. over 250 maps for 23 species, totalizing nearly 40,000 identified spots, making it the biggest gel-based proteomics dataset accessible from a single interface. Here, we can select a 2DE map which will be displayed for inspection. The database can be queried by keywords (protein description, protein name, gene name, species, author, full text, protein spot serial number) or graphically by clicking on a spot. Each spot is linked to a page containing the corresponding gene (protein) information and identification details. Also, information is displayed about other spots in different maps, where product of the same gene is detected. All these spots are highlighted in the maps and the calculated parameters [isoelectric point (pI) and molecular weight (Mw)] are displayed. There is a possibility for cross-references and obtaining more information from different 2DE databases and from UniProtKB. UniProtKB, a comprehensive protein sequence knowledge base has two sections: UniProtKB/Swiss-Prot, which is manually curated and UniProtKB/TrEMBL that contains computer-annotated entries. UniProtKB/Swiss-Prot entries provide users with cross-links to about 100 external databases and with access to additional information or tools [52].
{"title":"Two-Dimensional Gel Electrophoresis as an Information Base for Human Proteome","authors":"S. Naryzhny","doi":"10.5772/INTECHOPEN.75125","DOIUrl":"https://doi.org/10.5772/INTECHOPEN.75125","url":null,"abstract":"The main intricacy in the human proteome is that it is tremendously complex and com- posed of diverse and heterogeneous gene products. These products are called protein species or proteoforms and are the smallest units of the proteome. In pursuit of the comprehensive profiling of the human proteome, significant advances should be performed. The approaches that allow disclosing and keeping the information about human proteome using two-dimensional gel electrophoresis (2DE) are described. Experimental identification methods such as mass spectrometry of high resolution and sensitiv- ity (MALDI-TOF MS and ESI LC-MS/MS) or immunodetection in combination with bioinformatics and 2DE can be used for the development of a comprehensive knowledge base of the human proteome. over 250 maps for 23 species, totalizing nearly 40,000 identified spots, making it the biggest gel-based proteomics dataset accessible from a single interface. Here, we can select a 2DE map which will be displayed for inspection. The database can be queried by keywords (protein description, protein name, gene name, species, author, full text, protein spot serial number) or graphically by clicking on a spot. Each spot is linked to a page containing the corresponding gene (protein) information and identification details. Also, information is displayed about other spots in different maps, where product of the same gene is detected. All these spots are highlighted in the maps and the calculated parameters [isoelectric point (pI) and molecular weight (Mw)] are displayed. There is a possibility for cross-references and obtaining more information from different 2DE databases and from UniProtKB. UniProtKB, a comprehensive protein sequence knowledge base has two sections: UniProtKB/Swiss-Prot, which is manually curated and UniProtKB/TrEMBL that contains computer-annotated entries. UniProtKB/Swiss-Prot entries provide users with cross-links to about 100 external databases and with access to additional information or tools [52].","PeriodicalId":186044,"journal":{"name":"Electrophoresis - Life Sciences Practical Applications","volume":"19 1","pages":"0"},"PeriodicalIF":0.0,"publicationDate":"2018-09-12","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"127816970","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}
Pub Date : 2018-03-28DOI: 10.5772/INTECHOPEN.75042
S. Viglio, Maddalena Cagnone, L. Chiarelli, R. Salvini, P. Iadarola
The current chapter was designed to keep the reader informed about the present status of pulmonary proteome. Taken together, the results documented here demonstrate that, after a decade of activity, proteomics of pulmonary diseases is catching up with its promise. The constantly growing number of reports in this area supports the view of this approach as one of the decisive methodological tools for the identification/characterization of diseaseassociated proteins. In terms of experimental procedures, the basic options available for proteomic investigations consist in the identification of proteins through the use of gelbased or gel-free techniques followed by MS. Obviously, the question arises of whether sophisticated technologies (such as the non-gel-based proteomic procedures) may currently be more fruitful, in terms of candidate protein marker identification, than “conventional” (read electrokinetic) approaches. In light of the versatility and high degree of reproducibility shown by these new potent strategies, a positive answer is perhaps not surprising. Nevertheless, as documented in this chapter, despite being less sophisticated than competing ones, gel-based techniques still represent a widely used procedure able to generate a reliable protein “fingerprint” and to produce qualitative and quantitative information on the protein patterns of a variety of human fluids.
{"title":"The Role of One- and Two-Dimensional Electrophoretic Techniques in Proteomics of the Lung","authors":"S. Viglio, Maddalena Cagnone, L. Chiarelli, R. Salvini, P. Iadarola","doi":"10.5772/INTECHOPEN.75042","DOIUrl":"https://doi.org/10.5772/INTECHOPEN.75042","url":null,"abstract":"The current chapter was designed to keep the reader informed about the present status of pulmonary proteome. Taken together, the results documented here demonstrate that, after a decade of activity, proteomics of pulmonary diseases is catching up with its promise. The constantly growing number of reports in this area supports the view of this approach as one of the decisive methodological tools for the identification/characterization of diseaseassociated proteins. In terms of experimental procedures, the basic options available for proteomic investigations consist in the identification of proteins through the use of gelbased or gel-free techniques followed by MS. Obviously, the question arises of whether sophisticated technologies (such as the non-gel-based proteomic procedures) may currently be more fruitful, in terms of candidate protein marker identification, than “conventional” (read electrokinetic) approaches. In light of the versatility and high degree of reproducibility shown by these new potent strategies, a positive answer is perhaps not surprising. Nevertheless, as documented in this chapter, despite being less sophisticated than competing ones, gel-based techniques still represent a widely used procedure able to generate a reliable protein “fingerprint” and to produce qualitative and quantitative information on the protein patterns of a variety of human fluids.","PeriodicalId":186044,"journal":{"name":"Electrophoresis - Life Sciences Practical Applications","volume":"30 1","pages":"0"},"PeriodicalIF":0.0,"publicationDate":"2018-03-28","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"134092402","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}
Pub Date : 2018-03-14DOI: 10.5772/INTECHOPEN.74925
Niu-Jin Tan, L. J. Daim, A. A. Jamil, N. Mohtarrudin, Karuppiah Thilakavathy
Spontaneous unexplained preterm labor with intact membrane (sPTL-IM) remains as an unresolved challenge in obstetrics due to the complex syndromes involved during preterm birth. Two dimensional-gel electrophoresis (2D-GE) coupled with matrix-assisted laser desorption/ionization-time of flight/time of flight (MALDI TOF/TOF) mass spectrometry has become an alternative in screening for potential novel protein-based biomarkers and revealing the pathophysiology of sPTL-IM. To achieve this objective, protein extracted from fetal and maternal sides of the placenta obtained from sPTL-IM (n = 5) and the respective control (n = 10) groups were separated and compared using 2D-gel electrophoresis. MALDI-TOF/TOF mass spectrometry was utilized to identify the differentially expressed proteins between both groups, and the molecular functions of these proteins were studied. A total of 12 proteins were significantly differentiated in sPTL-IM over the control. Differentially expressed proteins were identified to have involved in structural/cytoskeletal components, immune responses, fetal and placenta development, and anticoagulation cascade. More proteins were found to be differentially expressed in the fetal side compared to the maternal side of the placenta. This postulates that the influence of sPTL-IM from fetus is greater than that of the mother. Ultimately, these results might lead to further investigations in elucidating the potential of these proteins as biomarkers and/or drug targets.
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