Kinetic Analysis of Bacteriophage Sf6 Binding to Outer Membrane Protein A Using Whole Virions

Natalia B. Hubbs; Mareena M. Whisby-Pitts; Jonathan L. McMurry

Our previous work showed that OmpA surface loops mediate Sf6 infection and confer host range [23] . Individual amino acid substitutions in OmpA loops result in a range of Sf6 infection efficiencies [23] . In an effort to better understand Sf6 interactions with OmpA, and how variations affect binding, we used biolayer interferometry (BLI) to determine the kinetics of these interactions. BLI is an optical biosensing technique used to measure the kinetic parameters of biomolecular interactions [28, 29] . It works by tethering one binding partner (the ligand, in this study, whole phage) to a fiber optic sensor tip. The ligand-loaded sensor is then dipped into a sample that contains a known concentration of the binding partner (the analyte, in this study, purified receptor proteins). White light reflects off of two optical layers in the tip, establishing an interference pattern, which is measured by a photodetector. Binding between ligand and analyte causes the distance between optical layers to increase, resulting in a shift in the interference pattern. The shift (in nm) plotted against time in an association-then-dissociation experiment All rights reserved. No reuse allowed without permission.