Sedimentation coefficients molecules

At first glance, the contents of Chap. 9 read like a catchall for unrelated topics. In it we examine the intrinsic viscosity of polymer solutions, the diffusion coefficient, the sedimentation coefficient, sedimentation equilibrium, and gel permeation chromatography. While all of these techniques can be related in one way or another to the molecular weight of the polymer, the more fundamental unifying principle which connects these topics is their common dependence on the spatial extension of the molecules. The radius of gyration is the parameter of interest in this context, and the intrinsic viscosity in particular can be interpreted to give a value for this important quantity. The experimental techniques discussed in Chap. 9 have been used extensively in the study of biopolymers. 

In a solution of molecules of uniform molecular weight, all particles settle with the same value of v. If diffusion is ignored, a sharp boundary forms between the top portion of the cell, which has been swept free of solute, and the bottom, which still contains solute. Figure 9.13a shows schematically how the concentration profile varies with time under these conditions. It is apparent that the Schlieren optical system described in the last section is ideally suited for measuring the displacement of this boundary with time. Since the velocity of the boundary and that of the particles are the same, the sedimentation coefficient is readily measured. 

The particle can be assumed to be spherical, in which case M/N can be replaced by (4/3)ttR P2, and f by 671770R- In this case the radius can be evaluated from the sedimentation coefficient s = 2R (p2 - p)/9t7o. Then, working in reverse, we can evaluate M and f from R. These quantities are called, respectively, the mass, friction factor, and radius of an equivalent sphere, a hypothetical spherical particle which settles at the same rate as the actual molecule. 

Ribosomes, the supramolecular assemblies where protein synthesis occurs, are about 65% RNA of the ribosomal RNA type. Ribosomal RNA (rRNA) molecules fold into characteristic secondary structures as a consequence of intramolecular hydrogen bond interactions (marginal figure). The different species of rRNA are generally referred to according to their sedimentation coefficients (see the Appendix to Chapter 5), which are a rough measure of their relative size (Table 11.2 and Figure 11.25). 

The type IV collagen molecule is comprised of the N-terminal collagenous 7S domain, which has a sedimentation coefficient of the central major collagenous domain with more than 20 interruptions... 

Figure 1.7 Structure of a ribosome. It is composed of two subunits the large 60S subunit has a mass of 2800 kDa and is composed of three RNA molecules and about 50 protein molecules. The smaller AOS subunit contains one RNA molecule plus around 30 protein molecules and has an aggregate mass of 1400 kDa. ( S is an abbreviabon for a Svedberg, the unit of the sedimentation coefficient. This is measured in an analybcal ultracentrifuge and is related to, but not simply proporbonal to, molecular mass.)...

Neurotrophic factors responsible for neuronal survival, dendritic proliferation, and the activation of the different neurotransmission systems are present in the central nervous system [CNS). The most well-known one is the NGF, a peptidergic complex of 140 kd and with a sedimentation coefficient of 7s. NGF has three subunits, a, p, and y. Subunit p is the active part of the molecule. Other neurotrophic factors [F. ffefti 1994) include 1) brain-derived neurotrophic factor [BDNF), 2) neurotrophin 3, 3) neurotrophin 4/5, and 4) ciliary neurotrophic factor. 

The structure of concentrated solutions of branched molecules has also received little attention. It is probable that the network formed by entangled branched molecules displays a topological structure which is different from that formed by linear chains. It is known that the sedimentation properties of branched synthetic polymers differ, especially in good solvents, from those of linear polymers. The concentration dependence of the sedimentation coefficient is relatively more pronounced for branched polymers than that for linear ones 20). 

Conformation in solution is indicated by the way in which the hydro-dynamic properties of the macromolecules change with change in molecular weight. From trends in the intrinsic viscosity, the sedimentation coefficient, or the diffusion constant with molecular weight we can learn something about the conformation of the molecule in solution (19,20). 

The fluid resistance experienced by a macromolecular solute moving in dilute solution depends on the shape and size of the molecule. A number of physical quantities have been introduced to express this. Typical ones are intrinsic viscosity [ry], limiting sedimentation coefficient s0, and limiting diffusion coefficient D0. The first is related to the rotation of the solute, while the last two are concerned with the translational motion of the solute. A wealth of theoretical and experimental information about these hydrodynamic quantities is already available for randomly coiled chains (40, 60). However, the corresponding information on non-randomly coiled polymers is as yet rather limited in number and in variety. 

To minimize the effects of concentration dependence, the sedimentation coefficient is usually determined for a particular system at several concentrations and then extrapolated to infinite dilution (S°). If, on the other hand, the diffusion coefficient is also measured at several concentrations and extrapolated to infinite dilution (D°), then values of S° and D° may be related through the frictional coefficient to determine the molecular weight of the solute molecules... 

B 4. An enzyme has a sedimentation coefficient of 3.5 S. When a substrate molecule is bound into the active site of the enzyme, the sedimentation coefficient decreases to 3.0 S. Explain this change. 

The ribosomes of Escherichia coli (M = 2.3 x 106 dalton) consist of two dissimilar subunits. The larger one is the 50 S unit containing 32 unique proteins (L1-L34) and one molecule each of 5 S and 23 S rRNA. The smaller subunit has a sedimentation coefficient of 30 S and contains one molecule 16 S rRNA and 21 unique proteins (S1-S21). The proteins of both subunits could be separated by RPC on a C 18 column (250 x 4.6 mm do = 30 nm dP = 10 pm) by a gradient water/2-propanol with 0.1 % trifluoroacetic acid in both solvents (Fig. 20). With 2-propanol, 15 proteins of the... 

In an SV experiment, the sedimentation and the diffusion forces that determine the net rate of movement of the solvent—solution boundary, stem from two intrinsic properties of the solute molecules, their sedimentation coefficient (s) and their diffusion coefficient (D). Whereas D depends predominantly on the shape of the solute particles, s depends both on its shape and on its mass. The diffusion coefficient, D, is defined as the ratio of the flux of molecules (Jx, moving in the direction x under diffusive forces) to the concentration gradient of the molecules (dc/dx). The dependence of D on the molecular shape stems from its relation to the frictional coefficient f ... 

The maxima of the (s ) versus 5 curve, therefore, denotes the sedimentation coefficient of the molecule, s. The presence of multiple peaks could indicate either the existence ofconformationally distinct ensembles or self-association of the RNA molecules or both. 

What is the velocity of the molecule described in the preceding problem when in a centrifugal field of 75,000 x gl Assume its density to be 1.40 g/cm3. What is its sedimentation coefficient ... 

Ribosomes are large macromolecular complexes whose components contain all the information necessary for self-assembly. The E. coli ribosome has a sedimentation coefficient of 70 S and consists of two subunits (50 S and 30 S) with a total mass of 2.8 x 106 Da and with 58 different components. Three of these components are RNA molecules that together comprise 65 percent of the mass and they act as a framework or template for the ordering of the different proteins. When the pure dissociated components are mixed together in the proper order under the correct conditions they spontaneously reassemble to form a fully active ribosome (Fig. 5-1). 

The absorption spectrum studies presented above merely reflect the electronic environment of the molecule and do not give specific information about the type of interaction. The data which must be accounted for in considering a physical mode for the binding process can be derived from several different approaches. Hydro-dynamic measurements on the DNA-drug complex are of interest, since Lerman58, S9 has established that an increase in the intrinsic viscosity of DNA and a decrease in the sedimentation coefficient of the polymer are two criteria for intercalation of ring systems between base pairs of a double-helical DNA. 

If the sample contains several components whose sedimentation coefficients are not very different, this will lead to the formation of unusually broad bands. If there are larger differences in the sedimentation coefficients, then separation into several bands occurs. Sedimentation velocity runs can be used to analyse molecular interactions. If a relatively small molecule (say a protein) in excess, binds to a larger molecule (e.g., DNA) then normally two bands will be formed a fast-moving band comprising the free and bound forms of the large molecule (DNA), and a slower band consisting of the excess of the smaller protein molecule (Figure 4-41). 

Figure 9-11. Sedimentation coefficients of various biological molecules, subcellular organelles, and organisms. (Courtesy of Beckman Instruments, Palo Alto, Calif.)...

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