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Gel electrophoresis

About: Gel electrophoresis is a research topic. Over the lifetime, 26026 publications have been published within this topic receiving 1113565 citations.


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TL;DR: BN-PAGE allows the preparation of mitochondrial protein complexes from gram amounts of plant tissue, as the procedure only requires milligram amounts of organelles and reveals novel results for their composition, molecular mass and stoichiometry.
Abstract: Recently a powerful electrophoresis method for the native preparation and characterization of the respiratory protein complexes of mitochondria from fungi and mammals has been developed, which employs Coomassie dyes to introduce charge shifts on proteins (Schagger and von Jagow (1991) Anal Biochem 199, 223–231) The procedure, which is called ‘blue native-polyacrylamide gel electrophoresis’ (BN-PAGE), was modified and introduced for the analysis of mitochondria from higher plants BN-PAGE of mitochondrial protein from potato allows the separation of nine distinct protein complexes between 100 and 1000 kDa and reveals novel results for their composition, molecular mass and stoichiometry For the first time soluble mitochondrial protein complexes, like the HSP60 complex (750 kDa) and a complex of 200 kDa, which includes a formate dehydrogenase, are analysed by BN-PAGE Complex I from potato (1000 kDa) is about 100 kDa larger than the corresponding enzyme from beef and can be resolved into more than 30 different subunits on a second gel dimension The F1F0 ATP synthase (580 kDa) and the cytochrome c oxidase (160 kDa) from potato seem to contain more subunits than hitherto reported Direct sequencing of subunits revealed that the F1 part of the F1F0 ATP synthase lacks the oligomycin sensitivity conferring protein (OSCP), which was reported to be present in F1 parts of dicotyledonous plants, but contains the ATPase inhibitory protein N-terminal sequences of 16 mitochondrial proteins were obtained, several of which are presented for the first time from a plant source BN-PAGE allows the preparation of mitochondrial protein complexes from gram amounts of plant tissue, as the procedure only requires milligram amounts of organelles This potential of BN-PAGE is demonstrated by the separation and characterization of the mitochondrial enzyme complexes from Arabidopsis thaliana Further analysis of organellar protein complexes by BN-PAGE will allow the generation of ‘protein maps’ from different tissues and developmental stages or from mutant plants

208 citations

Journal ArticleDOI
TL;DR: The reported results suggest that TGF- beta (TGF-beta 1) and T GF-beta 2 may have evolved from a common progenitor.
Abstract: Human type beta 2 transforming growth factor (hTGF-beta 2) was purified from tamoxifen-supplemented, serum-free medium conditioned by the human prostatic adenocarcinoma cell line PC-3. The purification of hTGF-beta 2 was monitored in a growth inhibition assay and was achieved by batch purification on methylsilyl-controlled pore glass, followed by gel permeation chromatography and reversed-phase high-performance liquid chromatography. The overall recovery of hTGF-beta 2 was 75% of the initial activity and yielded 22 micrograms of hTGF-beta 2/L of conditioned medium. The concentration of hTGF-beta 2 required for half-maximal inhibition of Mv 1 Lu mink lung epithelial cells (CCl-64) was approximately 5 pM when assayed in the presence of 10% fetal bovine serum. The purified hTGF-beta 2 has a molecular weight of 24,000 when analyzed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and consists of two disulfide-linked, apparently identical polypeptide chains, with a molecular weight of 13,000. The amino-terminal sequence of hTGF-beta 2 was determined. Alignment of the amino acid sequences of hTGF-beta 2 and hTGF-beta reveals statistically significant sequence homology. On the basis of the extensive amino acid sequence homology, we propose the term TGF-beta 2 for this newly isolated polypeptide. The reported results suggest that TGF-beta (TGF-beta 1) and TGF-beta 2 may have evolved from a common progenitor.

207 citations

Journal ArticleDOI
TL;DR: The CDGE technique was used to screen 32 breast carcinomas that had been analyzed by immunohistochemical methods for altered p53 protein levels and whose DNA had already been shown to have loss of heterozygosity for a chromosome 17p marker, and concluded that CDGE is a rapid and effective technique to screen for p53 mutations.
Abstract: At present, mutation of the p53 gene appears to be the most common genetic alteration found in human cancers. These mutations can occur within many different regions of the gene. We have developed a modification of denaturing gradient gel electrophoresis termed "constant denaturant gel electrophoresis" (CDGE), which provides a rapid and sensitive method to screen the four conserved regions within the p53 gene where the majority of p53 mutations have been reported. The sensitivity of CDGE was first tested with known p53 mutations in all four conserved regions. The CDGE technique was then used to screen 32 breast carcinomas that had been analyzed by immunohistochemical methods for altered p53 protein levels and whose DNA had already been shown to have loss of heterozygosity for a chromosome 17p marker. By immunostaining techniques, only 6 of the 32 tumors had elevated p53 expression. However, CDGE detected p53 mutations in 11 of the 32 tumors. DNA sequence analysis was performed to determine the nucleotide positions of these mutations in all 11 samples. Loss of heterozygosity for the pYNZ22 or p144D6 markers did not associate with either the loss of heterozygosity at the p53 locus or the mutations detected by CDGE. We conclude that CDGE is a rapid and effective technique to screen for p53 mutations.

207 citations

Journal ArticleDOI
TL;DR: Electrophoresis of plasma membrane preparations after solubilization with sodium dodecyl sulfate and reduction with beta-mercaptoethanol showed that a protein having a molecular weight of 130,000 was specifically labeled by the radioactive photosensitive insulin, suggesting that this protein may be the insulin receptor.

207 citations

Journal ArticleDOI
TL;DR: A procedure has been developed for the isolation of human Factor VII to apparent homogeneity as judged by the analytical disc electrophoretic system of Davis and by sodium dodecyl sulfate (SDS)-polyacrylamide gel electrophoresis.

207 citations


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Performance
Metrics
No. of papers in the topic in previous years
YearPapers
202364
2022116
2021108
2020104
2019120
2018147