Protein structures are stable and functional over a small range of environmental conditions. Outside this range the pattern of interactions that stabilizes the tertiary structure is disrupted and the molecule denatures—activity disappears as the molecule loses its structure. Denaturation may be caused by many factors, which include excessive temperature, change of pH, and detergents. Concentrated solutions of urea (8 mol per liter) have long been used by biochemists to denature proteins. Unlike heat and pH, urea does not cause the protein to precipitate. Physicochemical techniques have shown that in urea solution all of the higher levels of structure are lost and that polypeptide chains adopt random, changing conformations. Reagents such as urea that do this are called chaotropic reagents. If the urea is removed (by dialysis or simply by dilution), the protein refolds, regaining its structure and biological activity. This shows that the sequence of amino acids contains all of the information necessary to specify the final structure. The refolding of a urea-denatured protein cannot be random. Even a small protein with 100 amino acids would take some 1050 years to try out all of the structural conformations available. The fact that refolding does happen and happens on a time scale of seconds tells us that there must be a folding pathway, and the process is not random. Secondary structures may form first and act as folding units. In the cell folding is assisted by proteins called chaperones (page 221).
Medical Protein Folding Gone Awry—Mad Cow Disease Relevance Some years ago one of the dogmas of biology was overthrown—it was long be-9-1 lieved that diseases could only be transmitted by structures that contained nucleic acids, that is, by viruses or by microorganisms. It is now clear that a group of brain diseases can be transmitted by a protein. These are the diseases called spongiform encephalopathies: bovine spongiform encephalopathy in cows (mad cow disease), scrapie in sheep, and Creutzfeldt-Jacob disease and kuru in humans. These diseases are rare in humans but have recently increased in farm animals.
The infectious agent is called a prion and is a protein. It is coded for by a gene the animals have as part of their genome. In healthy individuals the gene is expressed in the cells of the nervous system and generates an innocuous protein called PrPc (prion-related protein cellular). PrPc has a globular region at the C-terminal end but the N-terminal region seems to be unstructured.
Sometimes the same polypeptide chain folds up differently with the disordered N-terminal section instead folding into a structure rich in f sheet. This is called PrPsc (prion-related protein scrapie). The disease arises because PrPsc can cause normal PrPc protein molecules to fold up into the abnormal form. This keeps happening and lumps of PrPsc form that damage nerve cells. The evidence is that it spread in cattle herds through food that contained recycled meat from infected animals. It is not yet known how one PrPsc molecule triggers the aberrant folding of a normal PrPc molecule.
Before the problem was fully comprehended, infected animals had been used for human food. There have been a number of cases in people that are thought to have come from eating contaminated beef—these are new variant Creutzfeldt-Jacob disease. It remains to be seen how many people will develop this fatal disease.
If a cell is immersed in a solution more dilute than its own cytosol, it tends to swell, as we have seen in the case of plant cells (page 53). On the other hand, cells immersed in concentrated solutions shrink. The overall strength of a solution is its osmolarity, and the movement of water into or out of cells because of solution strength is called osmosis. Many fish are exposed to changes in the salt concentration of the water around them, either because they swim from salt water to fresh and back (salmon, eels) or because they live in estuaries. The cells of these fish adjust the osmolarity of their cytosol to match the surrounding water by increasing the concentration of various small, easily synthesized molecules, one of which is urea. However, this introduces a further problem because high concentrations of urea are chaotropic—they cause cellular proteins to unfold.
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