High resolution two-dimensional electrophoresis of basic as well as acidic proteins

O'Farrell, Patricia Z.; Goodman, Howard M.; O’Farrell, Patrick H.

doi:10.1016/0092-8674(77)90176-3

Cited by 6,484 publications

(5,188 citation statements)

References 8 publications

Supporting

Mentioning

5,126

Contrasting

Unclassified

Order By: Relevance

“…Cells were then sonicated in TM buffer (pH 7AE4), and by centrifugation of the resultant extracts at 24 000 g for 40 min at 4°C, the membranes were pelleted. Proteins in the whole cell fraction were extracted with lysis buffer and assayed by SDS-PAGE (Laemmli 1970) or by two-dimensional gel electrophoresis (2D-PAGE) (O'Farrell 1975;Ames and Nikaido 1976), whereas proteins in the membrane fraction were assayed by modified SDS-PAGE containing 4 mol l -1 urea (Lundrigan and Earhart 1981). Gels were stained with either silver or Coomassie brilliant blue.…”

Section: Analyses Of Proteinsmentioning

confidence: 99%

Characterization of a cationic surfactant-resistant mutant isolated spontaneously from Escherichia coli

et al. 2002

View full text Add to dashboard Cite

Section: Analyses Of Proteinsmentioning

confidence: 99%

Characterization of a cationic surfactant-resistant mutant isolated spontaneously from Escherichia coli

et al. 2002

View full text Add to dashboard Cite

“…Two-dimensional gels were performed as originally described (O'Farrell, 1975;O'Farrell et al, 1977). Samples were prepared as follows.…”

Section: Two-dimensional Gel Electrophoresismentioning

confidence: 99%

Pkn9, a Ser/Thr protein kinase involved in the development of Myxococcus xanthus

Hanlon

Inouye

1997

Molecular Microbiology

View full text Add to dashboard Cite

SummaryThe Myxococcus xanthus gene, pkn9, encodes a protein that contains significant homology with eukaryotic Ser/ Thr protein kinases. The pkn9 gene was singled out of a previously identified family of kinase genes by amplification techniques that displayed differences in kinase gene expression during selected periods of the M. xanthus life cycle. Pkn9 was constitutively expressed during vegetative growth and upregulated during the aggregation stage of early development. It consists of 589 amino acids, and its N-terminal 394 residues show 38% identity with both Pkn1 and Pkn2 of M. xanthus. This region also shows 29, 25 and 29% identity with myosin light-chain kinase, protein kinase C, and cAMP-dependent protein kinase, respectively. A 22-residue hydrophobic transmembrane domain separates the kinase domain from the 173-residue C-terminal domain that resides on the outside of the inner membrane. The C-terminal domain contains two sets of tandem repeats of 13 and 10 residues which have no known function. When expressed in Escherichia coli under the T7 promoter, Pkn9 was found to be phosphorylated on serine and threonine residues. Disruption of the pkn9 kinase catalytic subdomains I-III by the insertion of a kanamycin-resistance gene resulted in slightly delayed, smaller and more-crowded fruiting bodies, while spore formation was normal. Total deletion of the pkn9 gene caused severely reduced progression through development resulting in light loose mounds that become slightly more compact over time. Development progressed further at the centre than at the edge of the spot, and spore formation was significantly reduced. Twodimensional gel analysis revealed that both the disruption and the deletion of pkn9 prevented the expression of five membrane proteins (KREP9-1-4). These results suggest that the loss of Pkn9 kinase activity caused altered fruiting-body formation, the absence of the KREP9 proteins in the membrane, and reduced spore production.

show abstract

“…TWO-dimensional gel electrophoresis using nonequilibrium pH gradient (NEPHG) in the first dimension was carried out according to O'Farrell et al (1977); the second dimension was performed in 18% acrylamide gels, using the gel and buffer system described by Thomas and Kornberg (1975). Before being applied to the first dimension separation , material of 100,000 x g pellets was first digested with 0.1 mg/ml pancreatic RNAase (Boehringer, Mannheim, FRG) for 15 min at 37°C in order to remove nucleic acids that might interfere with the migration of the proteins into the gel.…”

Section: Gel Electrophoresis Of Proteins and Immunoblottingmentioning

confidence: 99%

Ribocharin: a nuclear Mr 40,000 protein specific to precursor particles of the large ribosomal subunit

Hügle

Scheer

Franke

1985

Cell

View full text Add to dashboard Cite

SummaryUsing a monoclonal antibody (No-194) we have identified, in Xenopus laevis and other amphibia, an acidic protein of M, 40,000 (ribocharin) which is specifically associated with the granular component of the nucleolus and nucleoplasmic 65S particles. These particles contain the nuclear 28S rRNA and apparently represent the precursor to the large ribosomal subunit in nucleocytoplasmic transit. By immunoelectron microscopy ribocharin has been localized in the granular component of the nucleolus and in interchromatin granules. During mitosis ribocharin-containing particles are associated with surfaces of chromosomes and are recollected in the reconstituting nucleoli in late telophase. We suggest that ribocharin is a specific component of precursor particles of the large ribosomal subunit, which dissociates from the 65S particle before passage through the nuclear envelope, and is reutilized in ribosome biogenesis.

show abstract

High resolution two-dimensional electrophoresis of basic as well as acidic proteins

Cited by 6,484 publications

References 8 publications

Characterization of a cationic surfactant-resistant mutant isolated spontaneously from Escherichia coli

Characterization of a cationic surfactant-resistant mutant isolated spontaneously from Escherichia coli

Pkn9, a Ser/Thr protein kinase involved in the development of Myxococcus xanthus

Ribocharin: a nuclear Mr 40,000 protein specific to precursor particles of the large ribosomal subunit

Contact Info

Product

Resources

About