Steered Molecular Dynamics Simulations of a Type IV Pilus Probe Initial Stages of a Force-Induced Conformational Transition

Type IV pili are long, protein filaments built from a repeating subunit that protrudes from the surface of a wide variety of infectious bacteria. They are implicated in a vast array of functions, ranging from bacterial motility to microcolony formation to infection. One of the most well-studied type IV filaments is the gonococcal type IV pilus (GC-T4P) from Neisseria gonorrhoeae, the causative agent of gonorrhea. Cryo-electron microscopy has been used to construct a model of this filament, offering insights into the structure of type IV pili. In addition, experiments have demonstrated that GC-T4P can withstand very large tension forces, and transition to a force-induced conformation. However, the details of force-generation, and the atomic-level characteristics of the force-induced conformation, are unknown. Here, steered molecular dynamics (SMD) simulation was used to exert a force in silico on an 18 subunit segment of GC-T4P to address questions regarding the nature of the interactions that lead to the extraordinary strength of bacterial pili. SMD simulations revealed that the buried pilin α1 domains maintain hydrophobic contacts with one another within the core of the filament, leading to GC-T4P's structural stability. At the filament surface, gaps between pilin globular head domains in both the native and pulled states provide water accessible routes between the external environment and the interior of the filament, allowing water to access the pilin α1 domains as reported for VC-T4P in deuterium exchange experiments. Results were also compared to the experimentally observed force-induced conformation. In particular, an exposed amino acid sequence in the experimentally stretched filament was also found to become exposed during the SMD simulations, suggesting that initial stages of the force induced transition are well captured. Furthermore, a second sequence was shown to be initially hidden in the native filament and became exposed upon stretching.Author Summary: There are a large number of infectious bacteria that can be harmful to humans. Some bacterial infections are facilitated by long, tether-like filaments called type IV pili which extend from the surface of bacterial cells and attach to the surface of host cells. Type IV pilus filaments can grow to be many micrometers in length (bacterial cells themselves, on average, are only a couple of micrometers in length and half a micrometer in diameter), and can exert very large forces (up to 100,000 times the bodyweight of the bacteria). Because they extend from the surface of the cell, type IV pili are very good candidates for drug targeting. Computer simulation was used to exert forces on a segment of one of these filaments, in an effort to mimic the effects of tension that would be experienced by the pilus upon binding during infection. Regions of the filament that become exposed to the external environment in the pulled state were determined, in an attempt to identify amino acid sequences that could act as targets for drug design.