(518f) Directed Evolution of Influenza Hemagglutinin (HA) Reveals Novel pH-Sensing Mutants

Hemagglutinin (HA) is a viral fusion protein that undergoes an irreversible conformational change upon acidification to catalyze the fusion of endosomal and viral membranes.  Given the ubiquity of membrane fusion processes in nature, engineering a fusion protein for use in a drug or gene delivery vesicle, possibly promoting liposome fusion, would be a worthwhile endeavor.  We have employed directed evolution to identify novel mutants with activation pH across a range of 4.8 – 6.0; wild-type HA activates around pH 5.2. On the surface of transduced cells, a small number of engineered HA with altered pH sensing phenotypes can trigger activation of an excess of wild-type HA (or vice versa) by an autocatalytic mechanism, suggesting the ability to develop modular protein-based materials with separately tuned sensing and effector modules. Examination of library mutants has enabled us to identify individual amino acids responsible for phenotypic changes and thus provides additional structure-function insight.