Compact protein

Equation (8.97) shows that the second virial coefficient is a measure of the excluded volume of the solute according to the model we have considered. From the assumption that solute molecules come into surface contact in defining the excluded volume, it is apparent that this concept is easier to apply to, say, compact protein molecules in which hydrogen bonding and disulfide bridges maintain the tertiary structure (see Sec. 1.4) than to random coils. We shall return to the latter presently, but for now let us consider the application of Eq. (8.97) to a globular protein. This is the objective of the following example. 

Fig. 5. Protein folding. The unfolded polypeptide chain coUapses and assembles to form simple stmctural motifs such as -sheets and a-hehces by nucleation-condensation mechanisms involving the formation of hydrogen bonds and van der Waal s interactions. Small proteins (eg, chymotrypsin inhibitor 2) attain their final (tertiary) stmcture in this way. Larger proteins and multiple protein assembhes aggregate by recognition and docking of multiple domains (eg, -barrels, a-helix bundles), often displaying positive cooperativity. Many noncovalent interactions, including hydrogen bonding, van der Waal s and electrostatic interactions, and the hydrophobic effect are exploited to create the final, compact protein assembly. Further stmctural...

It is essential that the solution be sufficiently dilute to behave ideally, a condition which is difficult to meet in practice. Ordinarily the dilutions required are beyond those at which the concentration gradient measurement by the refractive index method may be applied with accuracy. Corrections for nonideality are particularly difficult to introduce in a satisfactory manner owing to the fact that nonideality terms depend on the molecular weight distribution, and the molecular weight distribution (as well as the concentration) varies over the length of the cell. Largely as a consequence of this circumstance, the sedimentation equilibrium method has been far less successful in application to random-coil polymers than to the comparatively compact proteins, for which deviations from ideality are much less severe. 

It is worth mentioning that the analytical approaches outlined here and currently used to treat relaxation data assume that the overall and local dynamics are not coupled. While this is a reasonable assumption for small, compact proteins, it might not be true for sys-... 

Histones are highly basic, small, compact proteins, with a high affinity for DNA. They occur naturally, attached to the DNA of cell nuclei by ionic linkages. Their classification is based on the relative amounts of lysine and arginine. The galactosylated, lysine-rich histone HI was found to be superior to the H2-H4 histones as a DNA carrier for liver gene delivery (53). 

For typical compact proteins this plot has a positive slope, as the hydrophilic residues on the outside of the dissolved protein have a higher scattering density than the hydrophobic residues on the inside. For casein sub-micelles, the slope is negative (Stothart and Cebula, 1982) (Figure 2). This seems surprising at first sight, but the sub-micelles are so highly hydrated that all the constituent protein... 

Fig. 15.1 Structure of pegfilgrastim. A relatively compact protein (filgrastim) is shown at the top of this molecule model. Despite having a similar molecular weight to filgrastim (18.8 kDa), the PEG moiety (20 kDa), shown at the bottom, is loosely hydrated and occupies a relatively large volume. (Adapted with permission from [12]).

Another important event contributing to the progress in this field was the development of reaction microcalorimetry, which has permitted direct measurement of heat effects involved with the transfer of hydrophobic substances from a nonpolar environment to water. These processes have been thought to mimic the unfolding of compact protein, structures. Prior to the development of direct calorimetric techniques, all information on the interaction of a hydrophobic substance with water was obtained from equilibrium studies. However, the results were limited in accuracy, particularly those properties that are obtained by consecutive temperature differentiation of the solubility, for example, the change in heat capacity. 

For compact proteins with molecular masses of greater than 10,000 and saturation of native structure by intramolecular hydrogen bonds of about 0.75 0.10 mole of bonds per mole of amino acid residues, the asymptotic values of enthalpy and entropy of the conformational transition, calculated per amino acid residue, amount to A%H(TX) = (6.25 0.2) kJ mol-1 and A 5(7 x) = (17.6 0.6) J K-1 mol-1. For some noncompact proteins (e.g., histones) or small globular proteins with molecular masses... 

Equation (9.1) allows us in principle to draw a spectrum that matches the entire spectrum obtained experimentally. This is correct only if the recorded spectrum originates from one fluorophore (i.e., Trp in solution) or from compact protein within a folded structure. However, when the experimental spectrum originates from two fluorophores (i.e., mixtures of tyrosine and tryptophan in solution) or from a disrupted protein that has two classes of Trp residues, the calculated spectrum using Equation (9.1) does not match the recorded spectrum. 

Hydrophobic effects are thus of practical interest. If we accept the goal of a simple, physical, molecularly valid explanation, then hydrophobic effects have also proved conceptually subtle. The reason is that hydrophobic phenomena are not tied directly to a simple dominating interaction as is the case for hydrophilic hydration of Na+, as an example. Instead hydrophobic effects are built up more collectively. In concert with this indirectness, hydrophobic effects are viewed as entropic interactions and exhibit counterintuitive temperature dependencies. An example is the cold denaturation of globular proteins. Though it is believed that hydrophobic effects stabilize compact protein structures and proteins denature when heated sufficiently, it now appears common for protein structures to unfold upon appropriate cooling. This entropic character of hydrophobic effects makes them more fascinating and more difficult. 

The parameters of the rotatory and translational diffusion of the macromolecule as a whole and those of intramolecular large-scale motions depend on the size and shape of the macromolecule, on its hydrodynamic permeability and on the possible solvation of the solvent. (It is known that the latter factor should be taken into account in the study of compact protein globules in water.)... 

A.26.9 Sedimentation analysis separates macromolecules by applying a force to them while they re in solution. Their different frictional coefficients lead to different rates of sedimentation. Mass, size, and shape. A long floppy protein will sediment slower than a similarly massed compact protein. 

Figure 2.48 Three-dimensional structure of myoglobin. (A) A ribbon diagram shows that the protein consists largely of a helices. (B) A space-filling model in the same orientation shows how tightly packed the folded protein is. Notice that the heme group is nestled into a crevice in the compact protein with only an edge exposed. One helix is blue to allow comparison of the two structural depictions. [Drawn from lA6N.pdb.]...

RFl and RF2 are compact proteins that in eukaryotes resemble a tRNA molecule. When bound to the ribosome, the proteins unfold to bridge the gap between the stop codon on the mRNA and the peptidyl transferase center on the SOS subunit. Although the precise mechanism of release is not known, the release factor may promote, assisted by the peptidyl transferase, a water molecule s attack on the ester linkage, freeing the polypeptide chain. The detached polypeptide leaves the ribosome. Transfer RNA and messenger RNA remain briefly attached to the 70S ribosome until the entire complex is dissociated in a GT F-dependent fashion in response to the binding of EF-G and another factor, called the ribosome release factor (RRF) (Figure 30.25). 

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