Combating bacterial infections

Certain pathogenic bacteria have adopted a unique ‘style’ of killing its victim by boring nanoscale holes into its cellular membrane.The pore-forming toxins (PFTs) released by these bacteria rapidly puncture the target cell membrane, and the cell leaks to death in a process known as apoptosis. Scientists across the globe have been actively working on designing various nanoscale (1-100 nanometres) blockers to plug these pores and prevent cell damage. Recently, a group of scientists from the Indian Institute of Science, Bengaluru, have suggested an effective nanoscale blocker made up of polyamido-amine (PAMAM) dendrimers, a synthetic polymer.

Professor Prabal K Maiti, from the Department of Physics and Centre for Condensed Matter Theory and Professor K G Ayappa, Professor at the Department of Chemical Engineering and Centre for Biosystems Science and Engineering, along with their two research scholars - Taraknath Mandal and Subbarao Kanchi, have successfully demonstrated the potential of PAMAM dendrimers in plugging the pores formed by PFTs released during bacterial infection. This work is part of a larger research initiative at IISc aimed at understanding the interactions of proteins and nanoparticles on biological membranes funded by the Department of Science and Technology, Government of India. The word ‘dendrimer’ refers to a molecule with tree-like branching and is highly regarded for its flexible properties. PAMAM is a type of star-shaped dendrimer with repetitive branched subunits of amide and amine functional groups.

Overcoming challenges
The study, investigated PAMAM dendrimer’s efficacy on blocking pores formed by CytolysinA (ClyA), a pore forming toxin produced by certain strains of E coli and other bacteria. ClyA has been rarely studied in the past due to the challenges in determining its crystal structure. But, what inspired the team to go against the tide? Professor Ayappa explains, “The crystal structure of ClyA has recently been discovered, allowing us for the first time to study and gain insight into the working of these toxins in molecular detail.” The team used a molecular dynamic simulation computational technique which predicts the position and velocities of atoms and molecules to demonstrate the pore gating ability of pH controlled PAMAM dendrimers.“This pH responsive behaviour makes it ideal for various applications like catalysis, lubricants to sensors and as nanoscale scaffolds for molecular imprinting,” says Prabal.

The pH of the surrounding solvent molecules is one of the determining factors which influences the protonation (addition of a proton, H+) and non-protonation (without H+) of the dendrimers. “While designing a specific dendrimer to effectively block the pore, both the size of the dendrimer molecule and its charge, are important factors. Since the pore formed by ClyA is predominantly negatively charged, we protonated the dendrimer (making it positively charged) to facilitate entry of the dendrimer into the bacterial pore channel,” explains Prof Ayappa.

The investigation compared the pore blocking efficacy of PAMAM in their protonated and non-protonated states. The findings revealed that the protonated dendrimers could expand their shapes to fit the size of the pore lumen, hence, effectively gating the passage of water and ions through the pore. On the other hand, the non-protonated counterparts allowed the passage of ions due to its inability to fill the gap completely. Previous studies have shown that PAMAM dendrimers block pores formed by other PFTs with about 45% efficiency without protonation. The current study achieved a whopping 91% efficiency in blocking the pores.

The professors at IISc note that antibiotic resistant bacterial strains have given rise to the so called ‘superbug’, which cannot be treated by known antibiotics, resulting in infections that are often fatal. This computer simulation study sheds light on a potential therapeutic strategy where, instead of targeting the bacteria with antibiotics, the nanometer sized pores formed by the bacteria are blocked by dendrimers, thereby preventing cell death and reducing the infection’s intensity. “Although this pore blocking strategy has the potential to treat a wide variety of pore mediated bacterial infections such as food poisoning, the eventual success would depend on extensive laboratory and clinical testing,” indicate Professors Prabal and Ayappa.