Master of Science, Miami University, 2020, Chemistry and Biochemistry

Holins are membrane proteins encoded by bacteriophages for promoting cell lysis by forming fatal "holes." In phage infections of gram-negative bacteria cell lysis occurs destroying the three layers of the cell envelope: cytoplasmic membrane, peptidoglycan layer, and outer membrane by Holin, Endolysin, and Spanin proteins respectively. Holins are the key determinant of the host lysis timing in phage infections. Canonical holins of phage lambda are of two types, actual lethal holin - S105, and antiholin - S107 as a result of the dual start motif of the Sλ gene that encodes them. Previously, the structural dynamics and topology of this system have not been studied using EPR spectroscopy in native membrane mimetics. In this study, SDSL-EPR spectroscopy is used to unravel structural dynamics and topology of the S105 in a micelle, liposome, and SMALPs environments. CW-EPR line shapes of selected MTSL labeled cysteine mutants across the three transmembrane domains and C-terminus of S105 were used to probe the dynamics of S105. CW-EPR power saturation experiments were used to determine the topology of different segments of S105 in a membrane utilizing the accessibility of paramagnetic relaxants O2 and NiEDDA. Successful insertion of S105 in membrane mimetics was confirmed by this study.

Committee: Gary Lorigan Ph.D. (Advisor); Carole Dabney-Smith Ph.D. (Committee Chair); Richard Page Ph.D. (Committee Member); Scott Hartley Ph.D. (Committee Member) Subjects: Biochemistry; Chemistry; Physical Chemistry