Mechanochemistry of gram-positive bacterial adhesins - towards the rational design of anti-invasive strategies

点击翻译按钮获取中文摘要

At the onset of an infection, bacteria attach to the host's tissue using long hair-like appendages, dubbed pili. These pili are built by the sequential concatenation of hundredths of pilin proteins, capped by a tip adhesin protein that directly interacts with ligands exposed in the target cell. Aiming to detach bacteria and prevent infection, the host responds with a battery of defense mechanisms-such as coughing or sneezing- that challenge the mechanical tether established by the bacterium, subjecting the individual pilin proteins to forces that would unfold any known protein. However, to overcome these challenges, bacteria have evolved unique chemical traits that confer their pilin proteins with exceptional mechanical properties allowing them to remain firmly attached despite these formidable stresses. For example, the tip pilins often contain a rare internal thioester bond that allegedly enables them to establish a covalent and long-lasting interaction with the host cells. Similarly, shaft pilin proteins are equipped with unique internal isopeptide bonds that provide these proteins with outstanding mechanical stability. For these reasons, pilin proteins are recognized virulence factors and have become an enticing target for developing new antibacterial drugs, particularly in light of the increasing threat of antibiotic-resistant bacteria. However, developing drugs that target pilin mechanics requires understanding how these proteins behave under large mechanical forces, which cannot be done with classic structural or biochemical techniques. This requires implementing new experimental methods to measure the mechanical properties of pilin proteins, providing the fundamental basis for the rational design of new antiadhesive compounds. Here, we propose a multiscale research program to unravel the mechanochemical properties of tip and shaft pilin proteins to develop anti-adhesive compounds that could obliterate their mechanical properties and prevent invasion. We will first use magnetic tweezers force spectroscopy to probe the mechanics of the thioester in the tip pilin of two bacterial pathogens, S. pyogenes (necrotizing fasciitis) and S. pneumoniae (pneumonia), designing peptides that block this bond. Second, we will study the shaft pilins of three bacteria containing isopeptide bonds-S. mutans (dental cavities), S. pneumoniae, and C. diphteria (diphteria)-and design mimicking peptides that block these isopeptide bonds. Finally, we will test these anti-adhesive strategies on living bacteria, evaluating how these pathogens attach under force when treated with the blocking peptides. Conducting this research will be possible thanks to the combined expertise of the lead and partner laboratories. The lead laboratory will bring its expertise in protein nanomechanics and magnetic tweezers force spectroscopy, providing a unique experimental approach to measure these proteins under force; the partner laboratory will contribute with its biochemistry expertise and experience working with bacteria. Overall, we will develop an innovative research program that will significantly contribute to our fundamental understanding of bacterial adhesion, further providing the ground basis for developing a novel generation of anti-bacterial drugs targeting pilin nanomechanics.