Accelerating chaperones
Advanced NMR techniques have allowed a US team to map the molecular chaperone Hsp70 in different states looking at individual amino acids within the protein as they respond to different sample conditions. Such insights could accelerate drug development for controlling chaperone behaviour a potentially important aspect of treating a wide range of diseases.
Molecular chaperones such as Hsp70s prevent proteins from misfolding within the cell's protein factory, the ribosome. Without proper folding, proteins will not adopt the necessary tertiary or quaternary structure in some cases and so molecular chaperone helpers are an essential part of protein production. The heat shock protein (Hsp) group of chaperones is particularly active in avoiding misfolding issues when tissue is exposed to heat, during a fever, for instance. Drugs that inhibit specific chaperones in specific disease states could lead to new ways to treat disease. As Gierasch explains, "New proteins emerge into a challenging environment. It's very crowded in the cell and it would be easy for them to get their sticky amino acid chains tangled and clumped together. Chaperones bind to them and help to avoid this aggregation, which is implicated in many pathologies such as neurodegenerative diseases. This role of chaperones has also heightened interest in using them therapeutically."
Anticancer targets
Molecular chaperones are potential drug targets with a particular interest to those looking to block cell proliferation in cancer where cancer cells recruit chaperones to facilitate their uncontrolled growth. Now, biochemist Lila Gierasch of the University of Massachusetts Amherst and her colleagues have used NMR spectroscopy to examine the functioning of Hsp70 by "trapping" this chaperone in action.
Writing in the journal Cell this month, the team explains how the new information regarding chaperone activity is important because rapidly dividing cells use a lot of Hsp70. "The saying is that cancer cells are addicted to Hsp70 because they rely on this chaperone for explosive new cell growth," explains Gierasch. "Cancer shifts our body's production of Hsp70 into high gear. If we can figure out a way to take that away from cancer cells, maybe we can stop the out-of-control tumour growth. To find a molecular way to inhibit Hsp70, you've got to know how it works and what it needs to function, so you can identify its vulnerabilities."
Chaperones cycle rapidly through states in which they are tightly bound to the folding protein and then loosely bound so that once the protein is ready it can be released quickly to carry out its job in the cell. The strength of binding is determined by whether adenosine triphosphate (ATP) or adenosine diphosphate (ADP) is bound. Hsp70s create a "holding pattern" to keep the newly formed protein ready for use and protected from stressors. In the loose state it is free to fold or be picked up by another chaperone and taken to the site in the cell where it will carry out its particular function.
Tickling molecules
Gierasch and colleagues already understood the nature of the Hsp70 structure in both the tight and the loose binding states, but not the intermediate state between the two. Identifying the nature of this state is essential to understanding how the chaperone works. NMR allowed Gierasch, postdoctoral fellows Anastasia Zhuravleva and Eugenia Clerico to access the in-between state of the bacterial Hsp70, DnaK, from Escherichia coli. By creating a mutant version of this Hsp70, the team could stop the cycling between tight and loose states allowing them to obtain a structure for the transient intermediate.
"And if you want to make a drug that controls the amount of Hsp70 available to a cell, our work points the way toward figuring out how to tickle the molecule so you can control its shape and its ability to bind to its client. We're not done, but we made a big leap," Gierasch adds. "We now have an idea of what the Hsp70 structure is when it is doing its job, which is extraordinarily important."
