Constant sedimentation

The sedimentation constant of a polymer at zero concentration, S0, may formally be written as  

In earlier work, following Flory (48), R was assumed proportional to the radius of gyration, and if this were so the relation  

So far, this method appears not to have been used as such, but sedimentation measurements have been combined with those of other kinds. Thus Moore and co-workers 136) assumed Eq. (9.3) and also that g =gi following Thurmond and Zimm (33) these assumptions led to the relation  

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Chase, A. M., and Langridge, R. (1960). The sedimentation constant and molecular weight of Cypridina luciferase. Arch. Biochem. Biophys. 88 294-297. 

The quantity on the left, which is determined by measuring the rate of movement of the sedimentation boundary in the ultracentrifuge operated at the constant angular velocity co, is called the sedimentation constant s. Thus... 

The mass of the sedimenting particle could be deduced from its rate of sedimentation at high dilution in a given field, i.e., from its sedimentation constant, if the frictional coefficient / could be determined independently. Hates of diffusion may be utilized to secure this necessary supplementary information, since the diffusion constant D depends also on the frictional coefficient. Thus ... 

The change in the sedimentation constant with concentration enters solely from the change in 1//, and it is customary therefore to extrapolate a plot of 1/s against c to infinite dilution. The results of sedimentation studies by Newman and Eirich on several polystyrene... 

Fig. 49.—Reciprocal of the sedimentation constant s plotted against concentration for two polystyrene fractions in chloroform. Molecular weights 13X10 and 5.5X10 for fractions 1 and 19, respectively. (Newman and Eirich. )...

Table XLII.—Sedimentation Constants fob Two Series op Polymer Fractions...

Experimental results are consistent with this relation, but inaccuracies in sedimentation constants preclude precise evaluation of the empirical exponent. Similarly, the diffusion constant at infinite dilution, given by... 

The effect of alkali treatment on molecular weight (compare with the case of the starch components) has been studied treating a 5% solution of rabbit-liver glycogen in 2 N sodium hydroxide, for 90 minutes at 100°, decreased the sedimentation constant (Sits X 1013) from 86 to 57 (that is, by 34%).237... 

In sedimentation velocity experiments, a dilute solution in high centrifugal field gets separated into a clear pure solvent layer and a solution. The initial clear boundary between the two then spreads because of diffusion. The rate of movement of boundary can be measured by suitable methods. Sedimentation constants is given by... 

Once the amino acid has been bound to its tRNA, it can pass to the next phase of protein synthesis, involving its interaction with mRNA, which takes place on the ribosome, a molecular machine of enormous complexity. The ribosome of E. coli is a ribonucleoprotein assembly of molecular weight 2700 kDa, and sedimentation constant of 70S9. It is made up of roughly two-thirds RNA and one-third protein, and can be separated into a small (30S) and a large (50S) subunit. The 30S subunit contains 21 proteins and one 16S RNA molecule, while the large subunit has 34 different proteins and two RNA molecules, one 23S and one 5S. Despite its size and complexity, the structure of both ribosomal subunits has been determined to atomic resolution (Figure 4.32), and very recently the atomic structure of the 70S ribosome has been determined at 2.8 A resolution (Selmer et al., 2006). 

Sedimentation- diffusion Absolute fairly fast Requires 3 separate determinations (diffusion coefficient, sedimentation constant and partial specific volume) <5... 

The rate of sedimentation is defined by the sedimentation constant 5, which is directly proportional to the polymer mass m, solution density p, and specific volume of the polymer V, and inversely proportional to the square of the angular velocity of rotation o), the distance from the center of rotation to the point of observation in the cell r, and the fractional coefficient /, which is inversely related to the diffusion coefficient D extrapolated to infinite dilution. These relationships are shown in the following equations in which (1 — Vp) is called the buoyancy factor since it determines the direction of macromolecular transport in the cell. 

The thus determined molecular weight is an apparent one since s and D depend on the polymer concentration. Therefore, extrapolation to concentration zero is required. The sedimentation coefficient obtained by extrapolating c —> 0 is called sedimentation constant Sn ... 

Here is a constant which depends on solvent and temperature. Thus the sedimentation constant can be calculated from they-axis intercept when the reciprocal of the sedimentation coefficient s determined at different polymer con-... 

The concentration dependence of sedimentation causes an additional reshaping of the boundary, called boundary sharpening. The faster moving material is at a higher concentration than the slower material, with the result that the boundary is progressively sharpened as the concentration increases that is, it shifts to the left. The dependence of sedimentation constant on concentration and on the concentration of other molecules around the sedimenting molecule, which causes the above effects, are too difficult to measure. In some systems with spherical macromolecules, approximate corrections have been made (5). The extrapolation of s to zero polymer concentration should eliminate the major part of these effects. 

Tung (106) has proposed the combination of sedimentation constant data with those from GPC, so as to obtain a distribution of branching, without fractionation being required. 

At pH 12, the disulfide and noncovalent bonds are both broken, and the monomer with a sedimentation constant of 1.45 Svedberg units is released. From frictional ratios, the monomer appears to exist as a coil with a diameter of 16 A and a length of 150 A. Analysis of the primary structure of K-casein (Loucheux-Lefebvre et al. 1978) suggests considerable secondary structure in the monomer. 23% a-helix, 31% /3-sheets, and 24% 0-turns. In contrast, other investigators, using several different approaches, obtained a-helix contents ranging from 0 to 20.8% (Bloomfield and Mead 1975). Circular dichroism spectra on the monomer indicated 14 and 31% for a-helix and / -sheet, respectively (Loucheux-Lefebvre et al 1978). An earlier study of the optical rotatory dispersion of the K-casein monomer yielded values for the a-helix content ranging from 2 to 16% (Herskovits 1966). 

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