Transporters are among the major components in the multidrug resistance armoury of pathogenic bacteria. Understanding the molecular architecture of these cell membrane-embedded machines paves the way for designing inhibitors against them, thereby helping in the treatment of diseases caused by these bacteria. In a new study, researchers led by Aravind Penmatsa in the Molecular Biophysics Unit solved the atomic-resolution structure of one such transporter, NorC, in complex with an Indian camelid antibody (ICab), where they saw that not only did the transporter get locked in a single state, the ICab effectively plugged the transporter like a “bottle-cork”, rendering it incapable of interacting with antibacterial compounds.
NorC is a transporter found in Staphylococcus aureus that helps provide resistance against broad-spectrum antibiotics like norfloxacin and moxifloxacin. Transporters like NorC are present within the cell membrane, which makes it difficult to extract them in their native shape. It is also challenging to coax the molecules into a periodic arrangement which helps in the formation of crystals needed for performing X-ray crystallography. The researchers extracted NorC in its native form using specialised detergent mixtures followed by using a single-domain antibody fragment obtained from an Indian camel (immunisation done at National Research Center on Camel, Bikaner) to help crystallise the complex of the ICab bound to NorC. Camels, llamas and sharks have unique single-domain antibodies that are much smaller compared to conventional antibodies and interact more effectively with deep pockets on the surface of antigens.
Upon solving the structure of the NorC-ICab complex using X-ray crystallography, the team found that the ICab inserted itself into the substrate-binding cavity of the transporter, effectively blocking its access to an antibacterial compound. Though transporters are molecular machines that need to shift their conformation to transport their substrates, the team found that the binding of ICab to NorC locked it into a single conformation.
These insights will significantly help formulate strategies to counter transporter-mediated antibiotic resistance in pathogenic bacteria, using tools like camelid antibodies. The structure of NorC and the conformation it has been solved in is also the first-of-its-kind, providing a model to study similar drug efflux transporters from other pathogenic bacteria.