The Role of Chaperones in Type III Secretion
Creators: name: University Of California Office Of The President
Publisher: name: University of California Office of the President
publisherIdentifier: https://ror.org/00dmfq477
publisherIdentifierScheme: ROR
Year: 2024

description: One of the hallmarks of advanced acquired immune deficiency syndrome (AIDS) is an increased susceptibility to bacterial pathogens. Many of these bacteria attack the host by using a syringe-like apparatus to inject toxin proteins (or “effectors”) directly into cells of the host immune system. This mechanism is known as the type III secretion system (TTSS), and our long-term goal is to understand the steps required to inject bacterial toxins through the TTSS. Prior to injection, some toxins are bound to another protein, known as a chaperone. Although these chaperones are not injected (they remain in the bacterium) they are required for the injection of their toxic partners. It is not known why these chaperones are necessary, but two hypotheses have been proposed. One idea suggests that chaperones unfold the toxins, enabling them to pass through the narrow syringe-like injection apparatus (toxin unfolding is analogous to unwinding a ball of yarn so that it can be passed through a drinking straw). Alternatively, another hypothesis proposes that TTSS chaperones act by directing their bound effectors to a component of the syringe, possibly by creating a shape that identifies the toxin as a molecule that must be injected. Both hypotheses are supported by indirect, conflicting studies. What is missing is a direct test of the hypotheses: this direct test would simply compare an intact toxin in its free and chaperone-bound states. One tool, known as “Nuclear Magnetic Resonance (NMR) spectroscopy,” is used to generate a type of protein fingerprint, revealing the protein’s unique shape. I propose to use NMR to test the two hypotheses explaining chaperone action. My specific aim is to determine how the toxin’s shape is changed when it binds to a chaperone. If the chaperone were found to make part of the toxin more flexible, this would provide direct evidence for the unfolding hypothesis. If instead the chaperone were found to stabilize a new shape on the toxin, this would support the hypothesis that the chaperone’s role is to direct the formation of a shape that helps the toxin to be injected. This shape could act by identifying the molecule as a toxin. Here, I present preliminary data that offer the first glimpse at an intact toxin. Already, we have observed that a feature of the toxin is changed when it binds to its chaperone. In this proposal, I outline a plan to identify the location of this new feature, and to determine the chaperone causes it to become more rigid or more flexible. This direct approach will discriminate between the alternative hypotheses proposed to explain the mode of action of TTSS chaperones, and will provide valuable insights into the fundamental mechanism by which toxins are injected. With the increased occurrence of multi-drug resistant bacteria it is critical to identify new drug targets. In the long-term, this project could lead to novel therapeutic strategies that target the TTSS chaperones. Because the chaperones are relied upon by many pathogens, drugs disrupting their normal function will likely have broad applications, significantly impacting the wellbeing of patients suffering from AIDS.