Analytical ultracentrifugation cells

Figure 13.9. Basic components of typical single- and double-sector analytical ultracentrifuge cells. The sector angle is the angle subtended at the center of rotation.

EXAMPLE 7.17 Ribosomal RNA in eukaryotes is actually four separate RNA species 28S RNA, 18S RNA, 5.8S RNA, and 5S RNA. (The S notation is related to how fast a molecule sediments in an analytical ultracentrifuge cell.) The larger the number, the faster the sedimentation rate. A globular protein of 100 kDa has an S value of 6S, but note that the relationship between size and S value is a sublinear one see Chap. 4. The 28S, 18S, and 5.8S rRNAs are transcribed as long precursor pre-rRNAs of size 45S. 

Fig. 3. (a) Schematic diagram of analytical ultracentrifuge cell containing a migrating lipoprotein species, (b) Representation of the concentration of this lipoprotein as a function of distance from the center of rotation, (c) Representation of refractive index gradient as a function of distance from the center of rotation. The Schlieren optical system of the Spinco model E ultracentrifuge actually provides this type of plot. 

The reference run, made in a given analytical ultracentrifuge cell using the reference salt solutions, provides a baseline tracing which represents the refractive index gradients due to redistribution of salt ions superimposed upon other gradients, such as compressional effects and cell assembly factors. 

An analytical ultracentrifugation method for determining the molecular mass, diffusion coefficient, and/or state of oligomerization of a macromolecule by conducting sedimentation conditions to establish an equilibrium distribution of the macromolecule from the meniscus to the bottom of the observation cell. 

Cultures of cells from healthy individuals and from patients with XPG are irradiated with ultraviolet light. The DNA is isolated and denatured, and the resulting single-stranded DNA is characterized by analytical ultracentrifugation. 

Acid phosphatase of S. aureus PS 55 is eluted from the cell surface by 1.0 M KC1 at pH 8.5. Gel filtration of this material gave a 44-fold purification. The protein seems homogeneous by gel filtration, starch block electrophoresis, and analytical ultracentrifugation with the weight of approximately 58,000 (12a). 

In this study, model latexes were diluted with distilled water to 1 percent solids by weight. Individual samples were adjusted with sodium hydroxide to various pH values from 3 to 13 and allowed to equilibrate 24 to 48 hours depending on the pH drift observed. Sedimentation coefficients were determined at 30°C with a Beckman Model E analytical ultracentrifuge using a single sector cell. 

Figure 4-39. Sector-shaped cells of an analytical ultracentrifuge.

Figure 9-15. Poly I,G-effected separation of the complementary strands of A phage DNA in preparative and analytical CsCl gradients. Upper trace represents the absorbance (260 nm) of the 4-drop (50 tl) fractions (total volume 2.5 ml) measured in a 20 /u.1 microcuvette (2 mm light path). Lower trace represents the microdensitometric tracing of the photograph of the same undiluted material banded in the analytical ultracentrifuge (4°C, 3 mm cell) with added density marker DNA (Cytophaga johnsonii, 1.6945 g/cm dashed line). Peak C contains the DNA strands C, which preferentially bind poly I, G the complementary strands W band under peak W. Symbols dN and NN indicate the positions (densities) of the denatured and native Acbj DNA, respectively. [From Z. Hradecna and W. Szybalski, Virology, 32 633 (1967).]...

Here we describe studies of the interaction of interleukin-6 (IL-6) with a soluble form of its cell surface receptor (sIL-6R). A procedure utilising a competition approach is presented which allows the determination of the equilibrium constant in solution thus avoiding any potential problems associated with deviation in kinetic characteristics upon surface immobilisation. In addition, binding characteristics of stable monomeric and dimeric forms of IL-6 are presented to demonstrate both the drastic influence of solute multivalency on kinetic and equilibrium properties and the importance of auxiliary techniques such as analytical ultracentrifugation for the interpretation of SPR data. 

Liu, S. and W. F. Stafford (1992). A Real-Time Video-Based Rayleigh Optical System for an Analytical Ultracentrifuge Allowing Imaging of the Entire Centrifuge Cell. Biophys. J. 61, A476, 2745. 

Figure 13.8. Analytical ultracentrifuge equipped with a UV-vis optical detection system. The sliding slit at the photomultiplier allows positional recording of the absorbance along the cell.

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