Optimization of the first dimension for separation by two-dimensional gel electrophoresis of basic proteins from human brain tissue (Proteomics 2004, vol. 4, issue 1, pp. 27-30)

Abstract To understand at a molecular level the basis of the normal and pathological genetic differences between individuals it is necessary to begin a detailed two-dimensional gel electrophoretic mapping of the proteins of the human brain in normal individuals and those with various genetic neurological disorders. The present study is an examination of the polypeptide patterns of extracts of the human brain made with 9 mol/L urea solution. Details of the technique and the nomenclature of the patterns obtained are presented. the gels are separated into 20 sub-sections, based on standards with known molecular masses and isoelectric points. Groups of polypeptides within these sub-sections are identified by a number and a letter; the individual proteins are identified by a number. Thus, protein 1 in subsection 8, group b, would be designated 8b: 1. Subsequent papers in this series identify many of these proteins; show which proteins belong to the cytosol, synaptosome, myelin, and other brain fractions; show how these patterns vary between normal individuals and those with different neurological and psychiatric conditions; examine the effect of severe gliosis; and present the results of non-equilibrium gel electrophoresis for the more basic proteins.

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A proteomic investigation of protein adsorption to cerebral microdialysis membranes in surgically treated intracerebral hemorrhage patients

10.21203/rs.3.rs-25981/v1 ◽

2020 ◽

Author(s):

Lovisa Tobieson ◽

Zita Czifra ◽

Karin Wåhlén ◽

Niklas Marklund ◽

Bijar Ghafouri

Keyword(s):

Intracerebral Hemorrhage ◽

Human Brain ◽

Protein Adsorption ◽

Brain Tissue ◽

Cerebral Microdialysis ◽

Invasive Technique ◽

Human Brain Tissue ◽

Protein Patterns ◽

Molecular Weight Range ◽

Two Dimensional Gel Electrophoresis

Abstract Background: Cerebral microdialysis (CMD) is a minimally invasive technique for sampling the interstitial fluid in human brain tissue. CMD allows monitoring the metabolic state of tissue, as well as sampling macromolecules such as proteins and peptides. Recovery of proteins or peptides can be hampered by their adsorption to the CMD membrane as has been previously shown in-vitro, however, protein adsorption to CMD membranes has not been characterized following implantation in human brain tissue. Methods: In this paper, we describe the pattern of proteins adsorbed to CMD membranes compared to that of the microdialysate and of cerebrospinal fluid (CSF). We retrieved CMD membranes from three surgically treated intracerebral hemorrhage (ICH) patients, and analyzed protein adsorption to the membranes using two-dimensional gel electrophoresis (2-DE) in combination with nano-liquid mass spectrometry. We compared the proteome profile of three compartments; the CMD membrane, the microdialysate and ventricular CSF collected at time of CMD removal. Results: We found unique protein patterns for each of the three compartments, particularly in the molecular weight range of 10–35 kDa. Conclusion: This study highlights the importance of analyzing the membranes in addition to the microdialysate when using CMD to sample proteins for biomarker investigation.

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Two-dimensional gel electrophoresis of human brain proteins. V. Non-equilibrium gel electrophoresis, with detection of a myelin basic protein mutation--MBL-Duarte.

Clinical Chemistry ◽

10.1093/clinchem/28.4.813 ◽

1982 ◽

Vol 28 (4) ◽

pp. 813-818 ◽

Cited By ~ 24

Author(s):

D E Comings ◽

A Pekkula-Flagan

Keyword(s):

Human Brain ◽

Myelin Basic Protein ◽

Gel Electrophoresis ◽

Basic Protein ◽

Myelin Proteins ◽

Two Dimensional Gel Electrophoresis ◽

Brain Proteins ◽

Basic Proteins ◽

A Charge ◽

Non Equilibrium

Abstract To examine the basic human brain proteins, we subjected 9 mmol/L urea extracts to non-equilibrium gel electrophoresis. The pattern observed differs distinctly from that with equilibrium gel electrophoresis. With this technique, the myelin proteins (myelin basic protein, proteolipids, and basic Wolfgram proteins) and many other unindentified major basic proteins can be demonstrated. The myelin basic proteins occur as two major polypeptides of different charge and slightly different molecular mass, indicating the action of at least two genes. The proteolipid proteins occur as a long series of charge isomers, suggesting multiple genes or extensive post-transcriptional modification. In one patient with schizophrenia, a charge-change mutation of the larger myelin basic protein (MBL) was observed and is termed "MBL-Duarte."

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Specific changes in the protein composition of rat liver in response to the peroxisome proliferators ciprofibrate, Wy-14,643 and di-(2-ethylhexyl)phthalate

Biochemical Journal ◽

10.1042/bj2270767 ◽

1985 ◽

Vol 227 (3) ◽

pp. 767-775 ◽

Cited By ~ 26

Author(s):

T Watanabe ◽

N D Lalwani ◽

J K Reddy

Keyword(s):

Gel Electrophoresis ◽

Peroxisome Proliferator ◽

Peroxisome Proliferation ◽

Peroxisome Proliferators ◽

Two Dimensional ◽

Two Dimensional Gel Electrophoresis ◽

Basic Proteins ◽

Acidic Proteins ◽

Liver Homogenates ◽

Ethylhexyl Phthalate

The hypolipidaemic agents ciprofibrate and Wy-14,643 ([4-chloro-6-(2,3-xylidino)-2-pyrimidinylthio]acetic acid) and the phthalate-ester plasticizer di-(2-ethylhexyl)-phthalate (DEHP), like other peroxisome proliferators, produce a significant hepatomegaly and induce the peroxisomal fatty acid beta-oxidation enzyme system together with profound proliferation of peroxisomes in hepatic parenchymal cells. Changes in the profile of liver proteins in rats following induction of peroxisome proliferation by ciprofibrate, Wy-14,643 and DEHP have been analysed by high-resolution two-dimensional gel electrophoresis. The proteins of whole liver homogenates from normal and peroxisome-proliferator-treated rats were separated by two-dimensional gel electrophoresis using isoelectric focusing for acidic proteins and nonequilibrium pH gradient electrophoresis for basic proteins. In the whole liver homogenates, the quantities of six proteins in acidic gels and six proteins in the basic gels increased following induction of peroxisome proliferation. Peroxisome proliferator administration caused a repression of three acidic proteins in the liver homogenates. By the immunoblot method using polyspecific antiserum against soluble peroxisomal proteins and monospecific antiserum against peroxisome proliferation associated Mr 80000 polypeptide (polypeptide PPA-80), the majority of basic proteins induced by these peroxisome proliferators appeared to be peroxisomal proteins. Polypeptide PPA-80 becomes the most abundant protein in the total liver homogenates of peroxisome-proliferator-treated rats. These results indicate that ciprofibrate, DEHP and Wy-14,643 induce marked changes in the profile of specific hepatic proteins and that some of these changes should serve as a baseline to identify a set of gene products that may assist in defining the specific ‘peroxisome proliferator domain’.

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Efficient solubilization buffers for two-dimensional gel electrophoresis of acidic and basic proteins extracted from wheat seeds

Biochimica et Biophysica Acta (BBA) - Proteins and Proteomics ◽

10.1016/j.bbapap.2005.10.003 ◽

2006 ◽

Vol 1764 (4) ◽

pp. 641-644 ◽

Cited By ~ 13

Author(s):

Gurusamy Chinnasamy ◽

Christof Rampitsch

Keyword(s):

Gel Electrophoresis ◽

Two Dimensional ◽

Two Dimensional Gel Electrophoresis ◽

Basic Proteins ◽

Wheat Seeds

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A proteomic investigation of protein adsorption to cerebral microdialysis membranes in surgically treated intracerebral hemorrhage patients

10.21203/rs.3.rs-19619/v1 ◽

2020 ◽

Author(s):

Lovisa Tobieson ◽

Zita Czifra ◽

Karin Wåhlén ◽

Niklas Marklund ◽

Bijar Ghafouri

Keyword(s):

Intracerebral Hemorrhage ◽

Human Brain ◽

Protein Adsorption ◽

Brain Tissue ◽

Cerebral Microdialysis ◽

Invasive Technique ◽

Human Brain Tissue ◽

Protein Patterns ◽

Molecular Weight Range ◽

Two Dimensional Gel Electrophoresis

Abstract Background Cerebral microdialysis (CMD) is a minimally invasive technique for sampling the interstitial fluid in human brain tissue. CMD allows monitoring the metabolic state of tissue, as well as sampling macromolecules such as proteins and peptides. Recovery of proteins or peptides can be hampered by their adsorption to the CMD membrane as has been previously shown in-vitro, however, protein adsorption to CMD membranes has not been characterized following implantation in human brain tissue. Methods In this paper, we describe the pattern of proteins adsorbed to CMD membranes compared to that of the microdialysate and of cerebrospinal fluid (CSF). We retrieved CMD membranes from three surgically treated intracerebral hemorrhage (ICH) patients, and analyzed protein adsorption to the membranes using two-dimensional gel electrophoresis (2-DE) in combination with nano-liquid mass spectrometry. We compared the proteome profile of three compartments; the CMD membrane, the microdialysate and ventricular CSF collected at time of CMD removal. Results We found unique protein patterns for each of the three compartments, particularly in the molecular weight range of 10-35 kDa. Conclusion This study highlights the importance of analyzing the membranes in addition to the microdialysate when using CMD to sample proteins for biomarker investigation.

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Two-dimensional gel electrophoresis of human brain proteins. III. Genetic and non-genetic variations in 145 brains.

Clinical Chemistry ◽

10.1093/clinchem/28.4.798 ◽

1982 ◽

Vol 28 (4) ◽

pp. 798-804 ◽

Cited By ~ 23

Author(s):

D E Comings

Keyword(s):

Human Brain ◽

Gel Electrophoresis ◽

Low Frequency ◽

Genetic Variations ◽

Genetic Mutations ◽

Two Dimensional ◽

Chi Square ◽

Two Dimensional Gel Electrophoresis ◽

Brain Proteins ◽

Chi Square Analysis

Abstract To determine the frequency of genetic mutations, polymorphisms, and non-genetic variation in the major human brain proteins, I examined, by equilibrium two-dimensional gel electrophoresis, 145 brains from patients dying of a wide variety of psychiatric, neurological, and non-neurological disorders. Of 176 polypeptides screened, there was one polymorphism of glial fibrillary acidic protein (GFAP-Duarte). Chi square analysis indicated it was non-randomly distributed among different diseases. A possible mutation associated with Joseph's disease is being further investigated. Three examples of a possible mutation of protein 8c:1 were noted. No other genetic mutations were observed. This low frequency of polymorphisms is consistent with results for two-dimensional gel analysis of other tissue and species. The numerous non-genetic variations are described.

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