Small-Scale Density Gradient Sedimentation to Separate and Analyze Multiprotein Complexes

Methods ◽  
1997 ◽  
Vol 12 (3) ◽  
pp. 224-234 ◽  
Author(s):  
Naoko Tanese
Keyword(s):  
Density Gradient ◽  
Small Scale ◽  
Multiprotein Complexes ◽  
Density Gradient Sedimentation

scholarly journals Electron microscopy of polyribosomes synthesizing immunoglobin chains. Comparison with sizes determined by density-gradient-sedimentation data

1970 ◽  
Vol 118 (3) ◽  
pp. 385-389 ◽  
Author(s):  
S. de Petris
Keyword(s):  
Electron Microscopy ◽  
Electron Microscope ◽  
Size Distribution ◽  
Density Gradient ◽  
Plasma Cell ◽  
Sucrose Gradient ◽  
Cell Tumour ◽  
Density Gradient Sedimentation

Polyribosomes from the immunoglobulin-producing plasma-cell tumour X5563 were fractionated on a linear sucrose gradient and fractions containing polyribosomes carrying heavy (H) and light (L) immunoglobulin chains were examined with the electron microscope. The size distribution of the polyribosomes in the various fractions was determined. The results are consistent with previous estimates by Williamson & Askonas (1967) of the size of the polyribosomes synthesizing immunoglobulin chains, despite some discrepancies in their calculations based on sedimentation data. In particular, polyribosomes of up to 18 ribosomes were present in the part of the gradient containing H-chain polyribosomes. The polyribosomal clusters appeared to be true linear aggregates of uniformly spaced ribosomes, arranged mostly in open or zig-zag configuration. The appearance of most of them does not seem to support a general helical model of the polyribosome.

scholarly journals Evidence for 24,25-dihydroxycholecalciferol receptors in long bones of newborn rats

1982 ◽  
Vol 204 (1) ◽  
pp. 31-36 ◽  
Author(s):  
D Sömjen ◽  
G J Sömjen ◽  
Y Weisman ◽  
I Binderman
Keyword(s):  
Density Gradient ◽  
Deae Cellulose ◽  
Sedimentation Coefficient ◽  
Sucrose Density Gradient ◽  
Macromolecular Complex ◽  
Long Bones ◽  
Newborn Rats ◽  
Sucrose Density Gradient Sedimentation ◽  
Density Gradient Sedimentation ◽  
Binding Component

Several reports have appeared that suggest that 24,25-dihydroxycholecalciferol has a possible biological role in bone formation. We have utilized competition studies, saturation analysis, sucrose-density-gradient sedimentation and DEAE-cellulose chromatography to demonstrate that long bones of vitamin D-depleted newborn rats contain cytoplasmic and possibly nuclear receptors that bind 24,25-dihydroxycholecalciferol with specificity and high affinity (Kd = 1.79 nM). Sucrose-density-gradient analysis of the cytoplasmic 24,25-dihydroxycholecalciferol-binding component showed a single binding macromolecule for 24,25-dihydroxycholecalciferol with a sedimentation coefficient of 3.1 S. DEAE-cellulose chromatography showed a [3H]24,25, dihydroxycholecalciferol-macromolecular complex that binds to DEAE-cellulose and elutes between 0.15 and 0.21 M-KCl. The finding of 24,25-dihydroxycholecalciferol receptors in long bones of newborn rats suggests a possible involvement of 24,25-dihydroxycholecalciferol in the metabolism of developing skeletal tissues.

scholarly journals Rev1, Rev3, or Rev7 siRNA Abolishes Ultraviolet Light-Induced Translesion Replication in HeLa Cells: A Comprehensive Study Using Alkaline Sucrose Density Gradient Sedimentation

2010 ◽  
Vol 2010 ◽  
pp. 1-12 ◽  
Author(s):  
Jun Takezawa ◽  
Yukio Ishimi ◽  
Naomi Aiba ◽  
Kouichi Yamada
Keyword(s):  
Density Gradient ◽  
Hela Cells ◽  
Uv Light ◽  
Sucrose Density Gradient ◽  
Detectable Effect ◽  
Main Role ◽  
Template Strand ◽  
Apparent Evidence ◽  
Sucrose Density Gradient Sedimentation ◽  
Density Gradient Sedimentation

When a replicative DNA polymerase stalls upon encountering a lesion on the template strand, it is relieved by other low-processivity polymerase(s), which insert nucleotide(s) opposite the lesion, extend by a few nucleotides, and dissociate from the 3′-OH. The replicative polymerase then resumes DNA synthesis. This process, termed translesion replication (TLS) or replicative bypass, may involve at least five different polymerases in mammals, although the participating polymerases and their roles have not been entirely characterized. Using siRNAs originally designed and an alkaline sucrose density gradient sedimentation technique, we verified the involvement of several polymerases in ultraviolet (UV) light-induced TLS in HeLa cells. First, siRNAs to Rev3 or Rev7 largely abolished UV-TLS, suggesting that these 2 gene products, which comprise Polζ, play a main role in mutagenic TLS. Second, Rev1-targeted siRNA also abrogated UV-TLS, indicating that Rev1 is also indispensable to mutagenic TLS. Third, Polη-targeted siRNA also prevented TLS to a greater extent than our expectations. Forth, although siRNA to Polι had no detectable effect, that to Polκ delayed UV-TLS. To our knowledge, this is the first study reporting apparent evidence for the participation of Polκ in UV-TLS.

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