Cells possess a large number of chaperones which make sure that proteins behave correctly and do not cause damage. Scientists at the Centre for Molecular Biology in Heidelberg are investigating the mechanisms used by the complex network of chaperones to control the proper folding of cellular proteins.
It was common knowledge that the amino acid chains of cellular proteins fold spontaneously into their specific three-dimensional structure due to the physical properties of the individual amino acids in their aqueous environment. The best-known experimental evidence for this belief is the enzyme ribonuclease (RNase): RNase denatures and becomes non-functional upon exposure to heat, for example, but spontaneously folds into a functional conformation when it is placed in an aqueous solution.
It is now known that spontaneous folding can only occur to small proteins such as RNase (bovine RNase A has a molecular weight of only 13.7 kDa), which is also an unusually stable protein. As a result of these properties, it was also the first protein whose tertiary structure was clarified. Larger and more complex proteins can only fold properly with the assistance of specialised proteins that are known as molecular chaperones.
At the beginning of the 1980s, Professor R. John Ellis (University of Warwick, UK), who focused on the biogenesis of chloroplasts, identified a protein (chaperone) that temporarily bound to a newly-synthesised polypeptide, thereby preventing the polypeptide from aggregating, and enabled it to fold into its proper tertiary structure. Since his discovery, Ellis and other researchers have discovered many other chaperones in all types of cells, including bacterial and human cells. When a protein is being synthesised, hydrophobic amino acids appear that must find other hydrophobic amino acids to associate with. However, there is the danger that they could associate with other proteins, leading to non-functional aggregates. The probability of aggregation is reinforced by the crowded conditions that prevail in cells (high concentration of macromolecules that is anything but an ideal aqueous solution). To avoid this problem, protein chaperones, such as those discovered by Ellis, stabilise newly-synthesised polypeptides while they fold into their proper structure and prevent them from forming aggregates.
The study of chaperones, a large heterogeneous group of proteins, has become a vast field of research. On 31st May 2011, Professor R. John Ellis, who retired in 1996, delivered the “Croonian Lecture”, the Royal Society’s premier lecture in the biological sciences. The Croonian Lecture was created in 1701 at the request of the widow of William Croone, one of the founding members of the Royal Society. Ellis’s Croonian Lecture was entitled “Molecular Chaperones: how cells stop proteins from misbehaving”.
The word “chaperone” can also mean escort and was previously used by the British and French upper classes to refer to adults who accompanied and supervised marriageable young women on social occasions to make sure that they behaved correctly and did not engage in inappropriate social interactions. The word figuratively derives from a hood worn by noble men and women during 15th century Italy (Quattrocento). Ellis adopted the word from Ron Laskey at Cambridge University who used the word for a protein that is involved in assembling nucleosomes (histone-DNA complex).
More than thirty different protein classes that act as chaperones are now known. The majority of these chaperone classes have most likely evolved independently from each other. Major research centres on heat shock proteins (HSP), proteins that were originally discovered as a result of their increased expression when cells are exposed to elevated temperatures.
In addition to temperature, factors such as oxidative stress or toxic substances are also known to trigger the synthesis of heat shock proteins (HSP). These proteins have the same function as proteins that function as chaperones during the biosynthesis of proteins: they prevent unwanted protein aggregation by recognising and binding to improperly folded amino acids and turning them back into their proper functional structure. HSPs are most likely found in all organisms. Their molecular interactions also involve other cellular components, including co-chaperones and proteases, with which they form a complex signalling network that is indispensible for the control of protein folding and for the regulation of protein homoeostasis in cells.Professor Dr. Bernd Bukau, Director of the Centre for Molecular Biology at the University of Heidelberg (ZMBH), and his team are investigating the molecular mechanisms of the network of chaperones and proteases that controls cellular protein folding using genetic, cell biological, biochemical and biophysical methods and model organisms such as bacteria, yeast and mammalian cells.
Heat shock proteins of the Hsp70 family (referred to as such as they have a molecular weight of 70 kDa) are key components of the cellular chaperone system that is ubiquitously expressed in bacteria and eukaryotes and that controls the proper folding and activation of numerous proteins. Hsp70 chaperones consist of an N-terminal ATPase domain and a C-terminal substrate-binding domain. The major effect of the Hsp70 machinery depends on the change of the molecule’s conformation due to its interaction with ATP. When Hsp70 binds to ATP, a groove with a peptide-binding site opens. When ATP is subsequently hydrolysed to ADP, the binding pocket closes, trapping the substrate peptide tightly. In cooperation with a group of researchers led by Dr. Matthias P. Mayer at the ZMBH, Bukau and his team performed detailed structure-function analyses to elucidate how the ATP- and ADP-bound states are linked to the change of conformation. Stress, i.e. elevated temperature, triggers the activation of the Sigma 32 heat shock transcription factor in bacteria. The ZMBH researchers have been able to show that the binding of DnaK (an isoform of Hsp70) increases the activity of Sigma 32. The co-chaperone DnaJ binds at a different site on Sigma 32, thereby triggering structural changes that facilitate DnaK binding and lead to the controlled degradation of Sigma 32. Bukau highlights that these findings are the first clear evidence of Hsp70-induced changes in substrate conformation. Other projects being carried out by Bukau’s group of researchers focus on the folding of newly-synthesised proteins at the bacterial ribosome which involves a chaperone known as “trigger factor”. They also are also investigating what are known as AAA+ chaperones that play a major role in regulated proteolysis and bacterial pathogenesis, and on strategies that enable cells to prevent the aggregation of proteins.