Harvard Catalyst Profiles

Antibiotic resistance is a major impediment to successful treatment of patients. Multidrug efflux pumps contribute to broad-spectrum antimicrobial resistance. This mechanism of resistance is particular important in certain pathogen groups like Pseudomonas, Acinetobacter, Burkholderia and Stenotrophomonas, sometimes leaving few if any treatment options. The goal of this student project is to understand how these efflux pumps function. Based on structural understanding of efflux pumps and cross comparisons among efflux pumps and the known efflux substrates, we will generate hypothesis about mechanisms of specific substrate recognition, and test these hypothesis by directed mutation of the pump amino acid sequence, and then measuring efflux of antibiotics of interest. This is a great project for a several month to a one year commitment as the questions are well defined and will allow the student to generate their own hypotheses and test them. Techniques would include highly efficient directed mutagenesis, structural predictions based on programs such as Pymol, Maestro, and alphafold2 to drive hypothesis generation and interpret results, and minimal inhibitory concentration analysis. We collaborate extensively with Ed Yu (Case Western Reserve University School of Medcine) who solved the first structure of a bacterial efflux pump, and is a leader in the structural biology of RND efflux pumps of interest. In collaboration with Dr. Yu, we also may also express and isolate the efflux pump proteins, and Dr. Yu's laboratory will solve the structures of the pumps by cryo-EM allowing us to confirm predicted hypotheses related to structure function correlations. Our long-term goal in underestanding, structure function correlations of efflux pumps is for future design of therapeutics that can inactivate pumps and thereby restore activity of existing antibiotics. Conversely, with a better understanding of pump substrate binding we can apply this knowledge to development of antibiotics that are no longer pump substrates and can then function in pump activated organisms. We are actively interested in design of such antibiotics. I have had other Harvard Medicla Students in the laboratory, and would welcome involvement of medical students. More information about he laboratory can be found at https://www.kirbylab.org