摘要:

AbstractNine previously uncharacterized membrane glycoproteins from normal rat liver have been analyzed by amino acid sequencing from two‐dimensional polyacrylamide gel electrophoresis (2‐D PAGE) after transblotting to Immobilon‐P membranes. Three of these components show altered levels of expression in liver tumors. A single electroblotted polyacrylamide gel yielded sufficient quantities of these glycoproteins for amino acid sequencing and theN‐terminal structure could be determined for four of them. The remaining five glycoproteins of interest were not sequenceable in this manner, presumably because they had blockedN‐termini. Prior to electrophresis, two enrichment methods were applied to the crude liver membrane preparations: affinity chromatography with concanavalin A to isolate the plasma membrane glycoproteins and then fast protein liquid chromatography on Superose 12 to obtain components having a specific range of molecular weights. These materials were next subjected to 2‐D PAGE using pH 4–6 carrier ampholytes in the first dimension and 7.5% sodium dodecyl sulfate gels in the second. The proteins were then electroblotted to Immobilon‐P membranes and located by staining with Coomassie Brilliant Blue R‐250. Our results demonstrate thatN‐terminal sequencing (gas‐phase) can be achieved on polypeptides obtained from approximately 250 μg of total glycoproteins appli

ISSN:0173-0835    

DOI:10.1002/elps.1150100702

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1989

数据来源: WILEY

摘要:

AbstractA soluble protein of unknown function was shown, by two‐dimensional polyacrylamide gel electrophoresis, to be present inNeisseria gonorrhoeaeat significantly higher concentrations than in the other related bacteria tested. The data indicate the possibility of this protein being specific toNeisseria gonorrhoeae. This protein was designated as NG 8.4 and purified. A radioimmunoassay was developed for the measurement of this protein and subsequently used to determine the degree of cross reaction exhibited by a number of species of bacteria. Of the bacteria tested only those of the genusNeisseriagave a significant reaction in the assa

ISSN:0173-0835    

DOI:10.1002/elps.1150100703

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1989

数据来源: WILEY

摘要:

AbstractProteins from nuclear plasma of mouse liver and brain and from the nuclear membranes of mouse liver were separated by two‐dimensional electrophoresis. For the purpose of comparison, liver cytosol proteins were also investigated. The protein samples were prepared from two inbred strains of the mouse (DBA/2J, C57BL/6J) and their hybrids. The patterns obtained were compared with regard to the composition and genetic variability (qualitative and quantitative variants) of proteins from different nuclear fractions and organs. The percentage (>30 %) of spots common to different organs (liver, brain), but from the same nuclear fraction (plasma) was greater than the percentage (<20 %) of spots common to different cell and nuclear fractions (cytosol, nuclear plasma and nuclear membranes) of the same organ (liver). Quantitative genetic variants occurred much more frequently than qualitative genetic variants (5.1 %vs.0.2 %; liver nuclear plasma). The incidence of genetic variants was much higher in liver (5.3 %) than in brains (0.0 %), and higher in solubilized nuclear proteins (5.3 %) than in structure‐bound nuclear proteins (2.1

ISSN:0173-0835    

DOI:10.1002/elps.1150100704

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1989

数据来源: WILEY

摘要:

AbstractFrom the cerebrospinal fluid of 32 patients with different neurological diseases immune complexes were isolated using protein A‐Sepharose. The isolated heavy and light chains and their constituents were analyzed by two‐dimensional gel electrophoresis. In addition to immunoglobulins, some proteins such as albumin, apolipo‐protein A‐I and a number of unknown proteins were detected in all preparations. A complex consisting of three proteins with molecular masses between 52–55 kDa reacted slightly with polyclonal antibodies to glial fibrillary acidic protein, Whether the linkage between these antigens and the Ig is due to the Fab region or the Fc region remains unknown in our study. In some immune complexes of neurological diseases such as amyotrophic lateral sclerosis, astrocytoma and multiple sclerosis, differences are easily recognizable in the ge

ISSN:0173-0835    

DOI:10.1002/elps.1150100705

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1989

数据来源: WILEY

摘要:

AbstractCerebrospinal fluid (CSF) proteins were separated into three main fractions by size exclusion‐high performance liquid chromatography (SE‐HPLC). Subsequent analysis of each fraction by two‐dimensional gel electrophoresis (2‐DE) facilitated the detection of trace components in CSF and additionally provided more information about the native properties of various proteins. Certain proteins are present in a polymeric form and appear in the high molecular weight SE‐HPLC fraction. In the middle molecular weight SE‐HPLC fraction we found a CSF‐specific transthyretinrelated protein by immunoblotting with polyclonal antibodies to transthyretin. Possible interpolypeptide disulfide bonds of such polymeric proteins were studied using a nonreducing 2‐DE system. This procedure revealed that all apolipoprotein E monomers in CSF, which are synthesized in astrocytes, are linked by disulfide bonds. In the CSF from a patient with clinically definite multiple sclerosis (MS), novel proteins appeared in the high molecular weight SE‐HPLC fraction, which are obscured by other proteins if tota

ISSN:0173-0835    

DOI:10.1002/elps.1150100706

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1989

数据来源: WILEY

摘要:

AbstractAnalysis of plasma proteins of Duchenne, muscular dystrophy (DMD) patients and normal age‐matched boys using two dimensional gel electrophoresis and densitometry indicated a three‐fold increase in the average haptoglobin level in DMD plasma. Two electrophoretic variants of haptoglobin in which the α2chain had more basic or more acidic spots were found in DMD patients with a rapidly progressing form of the disease. The basic variant was present in relatively low amount. Possible reasons for the elevated haptoglobin levels are discu

ISSN:0173-0835    

DOI:10.1002/elps.1150100707

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1989

数据来源: WILEY

摘要:

AbstractElectrophoretic light scattering has been used to investigate the interaction of ricin, a vegetal toxin, with cells. This technique allowed measurements in the presence of free ligand and proved particularly useful for the study of a system with low affinity. The electrophoretic mobility of erythrocytes and oligodendrocytes was found equal to 2.08 × 10−8and 2.35 × 10−8m2s−1V−1, respectively. Upon ricin binding, these values decreased significantly. This change was related to the saturation of the binding sites. The specificity of the interaction was demonstrated by conducting the experiments in the presence of lactose. This specific inhibitor fully prevented the ricin‐cell

ISSN:0173-0835    

DOI:10.1002/elps.1150100708

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1989

数据来源: WILEY

摘要:

AbstractFollowing polyacrylamide gel electrophoresis in an acetic/formic acid buffer, pH 2.0, in fabric reinforced gels, amino acids and oligopeptides could successfully be fixed in the gel by freeze‐drying. Lyophilization of the fabric reinforced polyacrylamide gel after electrophoresis resulted in a dry film which absorbed ninhydrin solution quickly and uniformly, thus improving the detection limit for amino acids and oligopeptides with molecular weights ranging from 189–1045. Most amino acids were detected with a sensitivity of 0.1–0.25 μg and for oligopeptides the detection limit was found to be 0.

ISSN:0173-0835    

DOI:10.1002/elps.1150100709

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1989

数据来源: WILEY

摘要:

AbstractFactor B is a glycoprotein which plays an essential role in the alternative pathway of complement activation. It carries the proteolytic activity of the convertases, and its physiological breakdown products Ba and Bb have some effects on the cells of the immune system. Human factor B exhibits a microheterogeneity and five isoforms are present in serum. The nature and origin of the microheterogeneity was investigated by using electrophoretic techniques. Treatments of factor B with neuraminidase and glycopeptidase F show that this microheterogeneity is mainly due to differences in its sialic acid content, varying from seven to eleven residues per molecule, and resulting in different oligosaccharide structures. However, deglycosylated factor B reveals a residual, nonallotypic variation in the Bb region of the polypeptide backbone. We confirm the presence of four asparagine‐linked oligosaccharide chains of the complex type in native factor B, two of which are located in the Ba fragment, and the two others in the Bb fragment. The prevalent isoform of the native protein carries two sialic acid residues per oligosaccharide chain. Biosynthesis experiments show that the microheterogeneity of secreted factor B from HepG2 cells is acquired during the processing of its glycans. However,in vitro‐secreted factor B is more heterogeneous than the serum protein. We propose a structural model for the microheterogeneity of the native protein and its physiological fragments. We discuss as well the feasibility of electrophoretic techniques to deal with microheterogeneity analy

ISSN:0173-0835    

DOI:10.1002/elps.1150100710

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1989

数据来源: WILEY

摘要:

AbstractThe polymorphism of alpha‐1‐antitrypsin (PI) has been studied by hybrid isoelectric focusing in miniaturized immobilized pH gradient gels, with an interelectrode distance of 55 mm, in two narrow ranges of pH 4.35–4.75 and 4.35–4.55, following rehydration with pH 4.2–4.9 carrier ampholytes. The use of the separatorN‐(2‐acetamido)‐2‐aminoethanesulfonic acid (ACES) in combination with carrier ampholytes for gel rehydration has been shown to enhance PI band sharpness. The influence of different additives (sucrose, sorbitol and glycerol) on the PI band pattern has also been evaluated. Glycerol has been shown to be responsible for the change in the relative mobility of the M3 band. The analysis of the minor M‐7 isoprotein zone by hybrid isoelectric focusing followed by silver staining has permitted a more reliable classification of PIM subtypes. A population study carried out with 164 unrelated individuals living in Spai

ISSN:0173-0835    

DOI:10.1002/elps.1150100711

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1989

数据来源: WILEY